1: Acta Crystallograph Sect F Struct Biol Cryst Commun. 2006 Dec 1;62(Pt
12):1286/9. Epub 2006 Nov 30.
Expression, purification, crystallization and preliminary X/ray analysis of
perakine reductase, a new member of the aldo/keto reductase enzyme superfamily
from higher plants.
Rosenthal C, Mueller U, Panjikar S, Sun L, Ruppert M, Zhao Y, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
Perakine reductase (PR) is a novel member of the aldo/keto reductase enzyme
superfamily from higher plants. PR from the plant Rauvolfia serpentina is
involved in the biosynthesis of monoterpenoid indole alkaloids by performing
NADPH/dependent reduction of perakine, yielding raucaffrinoline. However, PR can
also reduce cinnamic aldehyde and some of its derivatives. After heterologous
expression of a triple mutant of PR in Escherichia coli, crystals of the
purified and methylated enzyme were obtained by the hanging/drop
vapour/diffusion technique at 293 K with 100 mM sodium citrate pH 5.6 and 27%
PEG 4000 as precipitant. Crystals belong to space group C222(1) and diffract to
2.0 A, with unit/cell parameters a = 58.9, b = 93.0, c = 143.4 A.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 17142919 [PubMed / indexed for MEDLINE]
2: J Chromatogr Sci. 2006 Oct;44(9):557/60.
Quantitative determination of reserpine, ajmaline, and ajmalicine in Rauvolfia
serpentina by reversed/phase high/performance liquid chromatography.
Srivastava A, Tripathi AK, Pandey R, Verma RK, Gupta MM.
Analytical Chemistry Division, Central Institute of Medicinal and Aromatic
Plants, P.O./CIMAP, Lucknow 226015, India.
A sensitive and reproducible reversed/phase high/performance liquid
chromatography (HPLC) method using photodiode array detection is established for
the simultaneous quantitation of important root alkaloids of Rauvolfia
serpentina, namely, reserpine, ajmaline, and ajmalicine. A Chromolith
Performance RP/18e column (100 x 4.6/mm i.d.) and a binary gradient mobile phase
composed of 0.01 M (pH 3.5) phosphate buffer (NaH(2)PO(4)) containing 0.5%
glacial acetic acid and acetonitrile are used. Analysis is run at a flow rate of
1.0 mL/min with the detector operated at a wavelength of 254 nm. The calibration
curves are linear over a concentration range of 1/20 microg/mL (r = 1.000) for
all the alkaloids. The various other aspects of analysis (i.e., peak purity,
similarity, recovery, and repeatability) are also validated. For the three
components, the recoveries are found to be 98.27%, 97.03%, and 98.38%,
respectively. The limits of detection are 6, 4, and 8 microg/mL for ajmaline,
ajmalicine, and reserpine, respectively, and the limits of quantitation are 19,
12, and 23 microg/mL for ajmaline, ajmalicine, and reserpine, respectively. The
developed method is simple, reproducible, and easy to operate. It is useful for
the evaluation of R. serpentina.
PMID: 17059683 [PubMed / in process]
3: J Biosci Bioeng. 2006 Apr;101(4):287/96.
Metabolic engineering of cell cultures versus whole plant complexity in
production of bioactive monoterpene indole alkaloids: recent progress related to
old dilemma.
Pasquali G, Porto DD, Fett/Neto AG.
Programa de Pos/graduacao em Biologia Celular e Molecular, Centro de
Biotecnologia, Universidade Federal do Rio Grande do Sul, Av. Bento Goncalves
9500, Pr. 43.431, P.O. Box 15.005, CEP 91.501/970, Porto Alegre, RS, Brazil.
Monoterpene indole alkaloids (MIAs) are a large class of plant alkaloids with
significant pharmacological interest. The sustained production of MIAs at high
yields is an important goal in biotechnology. Intensive effort has been expended
toward the isolation, cloning, characterization and transgenic modulation of
genes involved in MIA biosynthesis and in the control of the expression of these
biosynthesis/related genes. At the same time, considerable progress has been
made in the detailed description of the subcellular/, cellular/, tissue/ and
organ/specific expressions of portions of the biosynthetic pathways leading to
the production of MIAs, revealing a complex picture of the transport of
biosynthetic intermediates among membrane compartments, cells and tissues. The
identification of the particular environmental and ontogenetic requirements for
maximum alkaloid yield in MIA/producing plants has been useful in improving the
supply of bioactive molecules. The search for new bioactive MIAs, particularly
in tropical and subtropical regions, is continuously increasing the arsenal for
therapeutic, industrially and agriculturally useful molecules. In this review we
focus on recent progress in the production of MIAs in transgenic cell cultures
and organs (with emphasis on Catharanthus roseus and Rauvolfia serpentina
alkaloids), advances in the understanding of in planta spatial/temporal
expression of MIA metabolic pathways, and on the identification of factors
capable of modulating bioactive alkaloid accumulation in nontransgenic
differentiated cultures and plants (with emphasis on new MIAs from Psychotria
species). The combined use of metabolic engineering and physiological modulation
in transgenic and wild/type plants, although not fully exploited to date, is
likely to provide the sustainable and rational supply of bioactive MIAs needed
for human well being.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 16716935 [PubMed / indexed for MEDLINE]
4: Plant Cell. 2006 Apr;18(4):907/20. Epub 2006 Mar 10.
The structure of Rauvolfia serpentina strictosidine synthase is a novel
six/bladed beta/propeller fold in plant proteins.
Ma X, Panjikar S, Koepke J, Loris E, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johanes
Gutenberg/University, D/55099 Mainz, Germany.
The enzyme strictosidine synthase (STR1) from the Indian medicinal plant
Rauvolfia serpentina is of primary importance for the biosynthetic pathway of
the indole alkaloid ajmaline. Moreover, STR1 initiates all biosynthetic pathways
leading to the entire monoterpenoid indole alkaloid family representing an
enormous structural variety of approximately 2000 compounds in higher plants.
The crystal structures of STR1 in complex with its natural substrates tryptamine
and secologanin provide structural understanding of the observed substrate
preference and identify residues lining the active site surface that contact the
substrates. STR1 catalyzes a Pictet/Spengler/type reaction and represents a
novel six/bladed beta/propeller fold in plant proteins. Structure/based sequence
alignment revealed a common repetitive sequence motif (three hydrophobic
residues are followed by a small residue and a hydrophilic residue), indicating
a possible evolutionary relationship between STR1 and several sequence/unrelated
six/bladed beta/propeller structures. Structural analysis and site/directed
mutagenesis experiments demonstrate the essential role of Glu/309 in catalysis.
The data will aid in deciphering the details of the reaction mechanism of STR1
as well as other members of this enzyme family.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 16531499 [PubMed / indexed for MEDLINE]
5: Acta Crystallograph Sect F Struct Biol Cryst Commun. 2006 Mar 1;62(Pt
3):257/60. Epub 2006 Feb 24.
Heterologous expression, purification, crystallization and preliminary X/ray
analysis of raucaffricine glucosidase, a plant enzyme specifically involved in
Rauvolfia alkaloid biosynthesis.
Ruppert M, Panjikar S, Barleben L, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
Raucaffricine glucosidase (RG) is an enzyme that is specifically involved in the
biosynthesis of indole alkaloids from the plant Rauvolfia serpentina. After
heterologous expression in Escherichia coli cells, crystals of RG were obtained
by the hanging/drop vapour/diffusion technique at 293 K with 0.3 M ammonium
sulfate, 0.1 M sodium acetate pH 4.6 buffer and 11% PEG 4000 as precipitant.
Crystals belong to space group I222 and diffract to 2.30 A, with unit/cell
parameters a = 102.8, b = 127.3, c = 215.8 A.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 16511316 [PubMed / indexed for MEDLINE]
6: Planta. 2005 Nov;222(5):888/98. Epub 2005 Aug 18.
Functional expression of an ajmaline pathway/specific esterase from Rauvolfia in
a novel plant/virus expression system.
Ruppert M, Woll J, Giritch A, Genady E, Ma X, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University, Staudinger Weg 5, 55128, Mainz, Germany.
Acetylajmalan esterase (AAE) plays an essential role in the late stage of
ajmaline biosynthesis. Based on the partial peptide sequences of AAE isolated
and purified from Rauvolfia cell suspensions, a full/length AAE cDNA clone was
isolated. The amino acid sequence of AAE has the highest level of identity of
40% to putative lipases known from the Arabidopsis thaliana genome project.
Based on the primary structure AAE is a new member of the GDSL lipase
superfamily. The expression in Escherichia coli failed although a wide range of
conditions were tested. With a novel virus/based plant expression system, it was
possible to express AAE functionally in leaves of Nicotiana benthamiana Domin.
An extraordinarily high enzyme activity was detected in the Nicotiana tissue,
which exceeded that in Rauvolfia serpentina (L.) Benth. ex Kurz cell suspension
cultures about 20/fold. This expression allowed molecular analysis of AAE for
the first time and increased the number of functionally expressed alkaloid genes
from Rauvolfia now to eight, and the number of ajmaline pathway/specific cDNAs
to a total of six.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 16133216 [PubMed / indexed for MEDLINE]
7: Acta Crystallogr D Biol Crystallogr. 2005 Jun;61(Pt 6):694/6. Epub 2005 May
26.
Crystallization and preliminary X/ray analysis of native and selenomethionyl
vinorine synthase from Rauvolfia serpentina.
Ma X, Koepke J, Bayer A, Fritzsch G, Michel H, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
Vinorine synthase (VS) is a central enzyme of the biosynthesis of the
antiarrhythmic drug ajmaline and is a member of the BAHD superfamily of
acyltransferases. So far, no three/dimensional structure with significant
sequence homology with VS is known. Crystals of VS and selenomethionyl/labelled
VS from the medicinal plant Rauvolfia serpentina have been obtained by the
hanging/drop technique at 305 K with ammonium sulfate and PEG 400 as
precipitants. VS crystals diffract to 2.8 A and belong to space group
P2(1)2(1)2(1), with unit/cell parameters a = 82.3, b = 89.6, c = 136.2 A. The
selenomethionyl VS crystal was nearly isomorphous with the VS crystal.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 15930622 [PubMed / indexed for MEDLINE]
8: Biochim Biophys Acta. 2004 Oct 1;1702(1):121/4.
Crystallization and preliminary X/ray crystallographic analysis of strictosidine
synthase from Rauvolfia: the first member of a novel enzyme family.
Ma X, Koepke J, Fritzsch G, Diem R, Kutchan TM, Michel H, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
Strictosidine synthase is a central enzyme involved in the biosynthesis of
almost all plant monoterpenoid indole alkaloids. Strictosidine synthase from
Rauvolfia serpentina was heterologously expressed in Escherichia coli. Crystals
of the purified recombinant enzyme have been obtained by the hanging/drop
technique at 303 K with potassium sodium tartrate tetrahydrate as precipitant.
The crystals belong to the space group R3 with cell dimensions of a=b=150.3 A
and c=122.4 A. Under cryoconditions (120 K), the crystals diffract to about 2.95
A.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 15450856 [PubMed / indexed for MEDLINE]
9: Biochim Biophys Acta. 2004 Sep 1;1701(1/2):129/32.
Vinorine synthase from Rauvolfia: the first example of crystallization and
preliminary X/ray diffraction analysis of an enzyme of the BAHD superfamily.
Ma X, Koepke J, Bayer A, Linhard V, Fritzsch G, Zhang B, Michel H, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
Crystals of vinorine synthase (VS) from medicinal plant Rauvolfia serpentina
expressed in Escherichia coli have been obtained by the hanging/drop technique
at 305 K with ammonium sulfate and PEG 400 as precipitants. The enzyme is
involved in the biosynthesis of the antiarrhythmic drug ajmaline and is a member
of the BAHD superfamily of acyltransferases. So far, no three/dimensional
structure of a member of this enzyme family is known. The crystals belong to the
space group P2(1)2(1)2(1) with cell dimensions of a=82.3 A, b=89.6 A and c=136.2
A. Under cryoconditions (120 K), a complete data set up to 2.8 A was collected
at a synchrotron source.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 15450182 [PubMed / indexed for MEDLINE]
10: Bioorg Med Chem. 2004 May 15;12(10):2787/95.
Acetyltransfer in natural product biosynthesis//functional cloning and molecular
analysis of vinorine synthase.
Bayer A, Ma X, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Institut fur Pharmazie, Johannes
Gutenberg/Universitat Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
Vinorine synthase (EC 2.3.1.160) catalyses the acetyl/CoA/ or CoA/dependent
reversible formation of the alkaloids vinorine (or 11/methoxy/vinorine) and
16/epi/vellosimine (or gardneral). The forward reaction leads to vinorine, which
is a direct biosynthetic precursor along the complex pathway to the
monoterpenoid indole alkaloid ajmaline, an antiarrhythmic drug from the Indian
medicinal plant Rauvolfia serpentina. Based on partial peptide sequences a cDNA
clone was isolated and functionally expressed in Escherichia coli. The Km values
of the native enzyme for gardneral and acetyl/CoA were determined to be 7.5 and
57 microM. The amino acid sequence of vinorine synthase has highest level of
identity (28/31%) to that of Papaver salutaridinol acetyltransferase, Fragaria
alcohol acyltransferase, and Catharanthus deacetylvindoline acetyltransferase
involved in morphine, flavor, and vindoline biosynthesis, respectively. Vinorine
synthase is a novel member of the BAHD superfamily of acyltransferases.
Site/directed mutagenesis of 13 amino acid residues provided clear evidence that
both, His160 and Asp164 of the consensus sequence HxxxD belong to the catalytic
center. The mutations also showed that an amino acid triad is not characteristic
of vinorine synthase. The experiments demonstrated the importance of the
conserved motif SxL/I/VD near the N/terminus and the consensus sequence DFGWG
near the C/terminal.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 15110860 [PubMed / indexed for MEDLINE]
11: Bioorg Med Chem. 2004 May 15;12(10):2781/6.
Purification and partial amino acid sequences of the enzyme vinorine synthase
involved in a crucial step of ajmaline biosynthesis.
Gerasimenko I, Ma X, Sheludko Y, Mentele R, Lottspeich F, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Institut fur Pharmazie, Johannes
Gutenberg/Universitat Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
The acetyl/CoA/dependent enzyme vinorine synthase was isolated from hybrid cell
suspension cultures of Rauvolfia serpentina and Rhazya stricta. The
sarpagan/type alkaloid gardneral was used as a substrate of the enzyme leading
to the ajmalan/type 10/methoxyvinorine. An HPLC/based assay was developed to
monitor vinorine synthase activity, which allowed establishing a five step
purification procedure combining anion exchange, hydrophobic interaction,
hydroxyapatite and gel filtration. Purification resulted in a yield of 0.2% and
an approximately 991/fold enrichment of the acetyltransfer activity. SDS/PAGE
analysis showed a Mr for the enzyme of approximately 50 kDa. The four peptide
fragments generated by proteolysis of the pure enzyme with endoproteinase LysC
and the N/terminal part of the enzyme were sequenced. The enzyme preparation (>
875/fold enrichment) delivering the N/terminal sequence was isolated from R.
serpentina cell suspensions. Sequence alignment of the five peptides showed
highest homologies in a range of 30/71% to acetyltransferases from other higher
plants involved in natural plant product biosynthesis. Based on the partial
sequences vinorine synthase is probably a novel member of the BAHD enzyme super
family.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 15110859 [PubMed / indexed for MEDLINE]
12: Bioorg Med Chem. 2003 Sep 1;11(18):3913/9.
In vivo monitoring of alkaloid metabolism in hybrid plant cell cultures by 2D
cryo/NMR without labelling.
Hinse C, Richter C, Provenzani A, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Institut fur Pharmazie, Johannes
Gutenberg/Universitat, Staudinger Weg 5, D/55099 Mainz, Germany.
Non/invasive measurements of alkaloid metabolism in plant cell suspension
cultures of a somatic hybrid from Rauvolfia serpentina Benth. ex Kurz and Rhazya
stricta Decaisne were carried out. When cell samples were taken sequentially
from a stock feeding experiment, measuring times for in vivo NMR of 40 min were
sufficient for following conversions of alkaloids at the natural abundance of
13C. Degradation of ajmaline added to the cells at 1.6 mM concentration to
raumacline could be monitored after 96 h on a standard 800 MHz NMR instrument
(Avance 800). Feeding vinorine an intermediate of ajmaline biosynthesis at 1.8
mM showed with a 500 MHz CryoProbe that the alkaloid enters two metabolic
routes. Vinorine is intracellularly transformed on route I through vellosimine
and 10/deoxysarpagine into sarpagine. On route II, the alkaloid is converted by
hydroxylation through vomilenine into the glucoside raucaffricine. Intracellular
alkaloid concentrations of approximately 500 microM are measurable in vivo with
cryogenic NMR technology.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12927851 [PubMed / indexed for MEDLINE]
13: Eur J Biochem. 2003 Feb;270(3):533/8.
Probing suggested catalytic domains of glycosyltransferases by site/directed
mutagenesis.
Hefner T, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Johannes Gutenberg/Universitat Mainz,
Germany.
The plant enzyme arbutin synthase isolated from cell suspension cultures of
Rauvolfia serpentina and heterologously expressed in Escherichia coli is a
member of the NRD1beta family of glycosyltransferases. This enzyme was used to
prove, by site/directed mutagenesis, suggested catalytic domains and reaction
mechanisms proposed for enzyme/catalyzed glycosylation. Replacement of amino
acids far from the NRD domain do not significantly affect arbutin synthase
activity. Exchange of amino acids at the NRD site leads to a decrease of
enzymatic activity, e.g. substitution of Glu368 by Asp. Glu368, which is a
conserved amino acid in glycosyltransferases located at position 2 and is
important for enzyme activity, does not serve as the nucleophile in the
catalytic centre as proposed. When it is replaced by Ala, the resulting mutant
enzyme E368A exhibits comparable activity as found for E368D in respect to
vanillin. Enzyme activities of wild/type and E368A towards several substrates
were not affected at the same level. His360 at position 1 of NRD1beta
glycosyltransferases occupies a more crucial role as expected. When it is
exchanged against other basic amino acids such as Lys or Arg the enzyme activity
decreases approximately 1000/fold. Replacement of His360 by Glu leads to a
mutant enzyme (H360E) with an approximately 4000/fold lower activity compared
with the wild/type. This mutein still produces a beta/glucoside, not an
alpha/glucoside and therefore indicates that generation of the typical E/E motif
of NRD1alpha glycosyltransferases does not convert a NRD1beta enzyme into a
NRD1alpha enzyme. The presented data do not support several suggestions made in
the literature about catalytic amino acids involved in the glycosyltransfer
reaction.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12542702 [PubMed / indexed for MEDLINE]
14: Phytochemistry. 2002 Nov;61(6):705/9.
Anhydronium bases from Rauvolfia serpentina.
Wachsmuth O, Matusch R.
Institut fur Pharmazeutische Chemie, Philipps/Universitat Marburg, Marbacher Weg
6, Germany.
Five anhydronium bases were isolated by preparative HPLC from a methanolic
extract of Rauvolfia serpentina roots. For the first time
3,4,5,6/tetradehydroyohimbine, 3,4,5,6/tetradehydro/(Z)/geissoschizol,
3,4,5,6/tetradehydrogeissoschizol and
3,4,5,6/tetradehydrogeissoschizine/17/O/beta/D/glucopyranoside were isolated
from a natural source. In addition, the well/known anhydronium base serpentine
was isolated. The structures of the compounds were determined by 1H and 13C NMR,
MS and UV.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12423893 [PubMed / indexed for MEDLINE]
15: J Chromatogr A. 2002 Aug 16;967(1):85/113.
High/performance liquid chromatographic, capillary electrophoretic and capillary
electrophoretic/electrospray ionisation mass spectrometric analysis of selected
alkaloid groups.
Stockigt J, Sheludk Y, Unger M, Gerasimenko I, Warzecha H, Stockigt D.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University, Mainz, Germany. stoeckig@mail.uni/mainz.de
Systems for efficient separation of selected alkaloid groups by high performance
liquid chromatography (HPLC), capillary electrophoresis (CE) and capillary
electrophoresis coupled with electrospray ionisation mass spectrometry
(CE/ESI/MS) are described. The optimized HPLC system was applied for the
separation of 23 standard indole alkaloids as well as for qualitative and
quantitative analyses of crude alkaloid extracts of Rauvolfia serpentina X
Rhazya stricta hybrid cell cultures. The developed conditions for CE analysis
proved to be efficient for separation of mixtures of standard indole and
beta/carboline alkaloids. The described buffer system is also applicable in the
combination of CE with electrospray ionisation mass spectrometry. This
analytical technique allowed the separation and identification of components of
standard indole alkaloid mixture as well as crude extracts of R. serpentina
roots, R. serpentina cell suspension cultures and cortex of Aspidosperma
quebracho/blanco. The influence of buffer composition and analyte structures on
separation is discussed.
Publication Types:
Research Support, Non/U.S. Gov't
Review
PMID: 12219932 [PubMed / indexed for MEDLINE]
16: J Nat Prod. 2002 Jul;65(7):1006/10.
New alkaloids of the sarpagine group from Rauvolfia serpentina hairy root
culture.
Sheludko Y, Gerasimenko I, Kolshorn H, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University, Staudinger Weg 5, D/55099 Mainz, Germany.
Three new monoterpenoid indole alkaloids, 19(S),20(R)/dihydroperaksine (1),
19(S),20(R)/dihydroperaksine/17/al (2), and
10/hydroxy/19(S),20(R)/dihydroperaksine (3), along with 16 known alkaloids 4/19
were isolated from hairy root culture of Rauvolfia serpentina, and their
structures were elucidated by 1D and 2D NMR analyses. Taking into account the
stereochemistry of the new alkaloids and results of preliminary enzymatical
studies, the putative biosynthetical relationships between the novel alkaloids
are discussed.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12141861 [PubMed / indexed for MEDLINE]
17: Planta Med. 2002 May;68(5):435/9.
Isolation and structure elucidation of a new indole alkaloid from Rauvolfia
serpentina hairy root culture: the first naturally occurring alkaloid of the
raumacline group.
Sheludko Y, Gerasimenko I, Kolshorn H, Stockigt J.
Institute of Pharmacy, Johannes Gutenberg/University, Mainz, Germany.
A new monoterpenoid indole alkaloid, 10/hydroxy/
N(alpha)/demethyl/19,20/dehydroraumacline ( 1), was isolated as a mixture of E/
and Z/isomers from hairy root culture of Rauvolfia serpentina Benth. ex Kurz
(Apocynaceae) and the structure was determined by 1D and 2D NMR analyses. The
new indole alkaloid represents the first naturally occurring alkaloid of the
raumacline group and its putative biosynthetical pathway is discussed.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12058321 [PubMed / indexed for MEDLINE]
18: Bioorg Med Chem. 2002 Aug;10(8):2479/83.
Deoxysarpagine hydroxylase//a novel enzyme closing a short side pathway of
alkaloid biosynthesis in Rauvolfia.
Yu B, Ruppert M, Stockigt J.
Johannes Gutenberg/University, Institute of Pharmacy, Department of
Pharmaceutical Biology, Staudinger Weg 5, 55099 Mainz, Germany.
Microsomal preparations from cell suspension cultures of the Indian plant
Rauvolfia serpentina catalyze the hydroxylation of deoxysarpagine under
formation of sarpagine. The newly discovered enzyme is dependent on NADPH and
oxygen. It can be inhibited by typical cytochrome P450 inhibitors such as
cytochrome c, ketoconazole, metyrapone, tetcyclacis and carbon monoxide. The
CO/effect is reversible with light (450 nm). The data indicate that
deoxysarpagine hydroxylase is a novel cytochrome P450/dependent monooxygenase. A
pH optimum of 8.0 and a temperature optimum of 35 degrees C were determined.
K(m) values were 25 microM for NADPH and 7.4 microM for deoxysarpagine.
Deoxysarpagine hydroxylase activity was stable in presence of 20% sucrose at /25
degrees C for >3 months. The analysis of presence of the hydroxylase in nine
cell cultures of seven different families indicates a very limited taxonomic
distribution of this enzyme.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12057637 [PubMed / indexed for MEDLINE]
19: Phytochemistry. 2002 Jul;60(5):467/74.
Increase of free cysteine and citric acid in plant cells exposed to cobalt ions.
Oven M, Grill E, Golan/Goldhirsh A, Kutchan TM, Zenk MH.
Lehrstuhl fur Botanik, Technische Universitat Munchen, Am Hochanger 4, D/85350,
Freising, Germany.
Cobalt complexation was investigated in a suspension cell culture of the cobalt
hyperaccumulator Crotalaria cobalticola. C. cobalticola cells were more tolerant
towards cobalt ions than the suspension cells of the non/accumulators Rauvolfia
serpentina and Silene cucubalus. While the concentration of various compounds
increased in cells of C. cobalticola challenged with cobalt ions, phytochelatin
biosynthesis was not induced. Instead, the exposure to cobalt ions resulted in
the increase of citrate and cysteine in cells. Size exclusion chromatography
demonstrated the co/elution of cobalt and cysteine in C. cobalticola cell
extracts. A significant increase in cysteine was observed also in cells of R.
serpentina and S. cucubalus when they were exposed to cobalt ions. These results
suggest that free cysteine is involved in cobalt ion complexation in plant
cells.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 12052512 [PubMed / indexed for MEDLINE]
20: Eur J Biochem. 2002 Apr;269(8):2204/13.
Heterologous expression of a Rauvolfia cDNA encoding strictosidine glucosidase,
a biosynthetic key to over 2000 monoterpenoid indole alkaloids.
Gerasimenko I, Sheludko Y, Ma X, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Institut fur Pharmazie, Johannes
Gutenberg/Universitat Mainz, Germany.
Strictosidine glucosidase (SG) is an enzyme that catalyses the second step in
the biosynthesis of various classes of monoterpenoid indole alkaloids. Based on
the comparison of cDNA sequences of SG from Catharanthus roseus and
raucaffricine glucosidase (RG) from Rauvolfia serpentina, primers for RT/PCR
were designed and the cDNA encoding SG was cloned from R. serpentina cell
suspension cultures. The active enzyme was expressed in Escherichia coli and
purified to homogeneity. Analysis of its deduced amino/acid sequence assigned
the SG from R. serpentina to family 1 of glycosyl hydrolases. In contrast to the
SG from C. roseus, the enzyme from R. serpentina is predicted to lack an
uncleavable N/terminal signal sequence, which is believed to direct proteins to
the endoplasmic reticulum. The temperature and pH optimum, enzyme kinetic
parameters and substrate specificity of the heterologously expressed SG were
studied and compared to those of the C. roseus enzyme, revealing some
differences between the two glucosidases. In vitro deglucosylation of
strictosidine by R. serpentina SG proceeds by the same mechanism as has been
shown for the C. roseus enzyme preparation. The reaction gives rise to the end
product cathenamine and involves 4,21/dehydrocorynantheine aldehyde as an
intermediate. The enzymatic hydrolysis of dolichantoside
(Nbeta/methylstrictosidine) leads to several products. One of them was
identified as a new compound, 3/isocorreantine A. From the data it can be
concluded that the divergence of the biosynthetic pathways leading to different
classes of indole alkaloids formed in R. serpentina and C. roseus cell
suspension cultures occurs at a later stage than strictosidine deglucosylation.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 11985599 [PubMed / indexed for MEDLINE]
21: Bioorg Med Chem. 2002 Jun;10(6):1913/8.
Vomilenine reductase//a novel enzyme catalyzing a crucial step in the
biosynthesis of the therapeutically applied antiarrhythmic alkaloid ajmaline.
von Schumann G, Gao S, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University, Staudinger Weg 5, D/55099, Mainz, Germany.
Delineation of the biochemical pathway leading to the antiarrhythmic Rauvolfia
alkaloid ajmaline has been an important target in biosynthetic research for many
years. The biosynthetic sequence starting with tryptamine and the monoterpene
secologanin consists of about 10 different steps. Most of the participating
enzymes have been detected and characterized previously, except those catalyzing
the reduction of the intermediate vomilenine. A novel NADPH/dependent enzyme
that reduces the intermediate has been isolated from Rauvolfia serpentina cell
suspension cultures. Vomilenine reductase (M(r )43 kDa, temp opt 30 degrees C,
pH opt 5.7/6.2), saturates the indolenine double bond of vomilenine with
stereospecific formation of 2beta(R)/1,2/dihydrovomilenine. The described
detection, enrichment and properties of the reductase not only closes a gap in
ajmaline biosynthesis but is also a prerequisite for overexpressing the protein
heterologously for final clarification of its molecular properties.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 11937349 [PubMed / indexed for MEDLINE]
22: Bioorg Med Chem. 2002 Jun;10(6):1731/41.
Arbutin synthase, a novel member of the NRD1beta glycosyltransferase family, is
a unique multifunctional enzyme converting various natural products and
xenobiotics.
Hefner T, Arend J, Warzecha H, Siems K, Stockigt J.
Johannes Gutenberg/University Mainz, Institute of Pharmacy, Department of
Pharmaceutical Biology, Staudinger Weg 5, D/55099, Mainz, Germany.
Plant glucosyltransferases (GTs) play a crucial role in natural product
biosynthesis and metabolization of xenobiotics. We expressed the arbutin
synthase (AS) cDNA from Rauvolfia serpentina cell suspension cultures in
Escherichia coli with a 6 x His tag and purified the active enzyme to
homogeneity. The recombinant enzyme had a temperature optimum of 50 degrees C
and showed two different pH optima (4.5 and 6.8 or 7.5, depending on the
buffer). Out of 74 natural and synthetic phenols and two cinnamyl alcohols
tested as substrates for the AS, 45 were accepted, covering a broad range of
structural features. Converting rates comparable to hydroquinone were not
achieved. In contrast to this broad acceptor substrate specificity, only
pyrimidine nucleotide activated glucose was tolerated as a donor substrate.
Nucleotide and amino acid sequence analysis revealed AS to be a new member of
the NRD1beta family of glycosyl transferases and placed the enzyme into the
group of plant secondary product GTs. Arbutin synthase is therefore the first
example of a broad spectrum multifunctional glucosyltransferase.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 11937332 [PubMed / indexed for MEDLINE]
23: Phytochem Anal. 2001 Mar/Apr;12(2):96/103.
Development of an efficient system for the separation of indole alkaloids by
high performance liquid chromatography and its applications.
Gerasimenko I, Sheludko Y, Unger M, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University, Staudinger Weg 5, D/55099 Mainz, Germany.
An efficient system for the analysis of indole alkaloids by HPLC on a
reversed/phase column using an ion pair technique is described. The optimised
chromatographic conditions allowed the successful separation of 22 standard
monoterpenoid indole alkaloids (including some isomers) and tryptamine. The
described HPLC system was applied to the analysis of alkaloids in intergeneric
somatic hybrid cell cultures of Rauvolfia serpentina x Rhazya stricta.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 11705246 [PubMed / indexed for MEDLINE]
24: Nat Prod Lett. 2001;15(2):119/24.
Glucosylation of isatin/3/oxime followed by 2D in situ NMR in plant cells at
highest magnetic field without labelling.
Hinse C, Unger M, Provenzani A, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, D/55099 Mainz, Germany.
The glucosylation of isatin/3/oxime (1) was monitored by in situ 2D 1H/13C
inverse correlated gradient assisted NMR spectroscopy in plant cell suspension
cultures of Rauvolfia serpentina without labelling. The applied high magnetic
field of 800 MHz allowed measurements within 20 min at concentrations of 1 of
5.76 mM. Complete glucosylation of 1 occurs inside the cells within 72 hours.
During this time isatin/3/oxime/glucoside (2) accumulates without further
metabolism.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 11561444 [PubMed / indexed for MEDLINE]
25: Biotechnol Bioeng. 2001 Sep;76(2):126/31.
Utilizing genetically engineered bacteria to produce plant/specific glucosides.
Arend J, Warzecha H, Hefner T, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University Mainz, Staudinger Weg 5, 55099 Mainz, Germany.
Plant/derived glucosides have attracted much attention due to their widespread
applications. This class of products is difficult to isolate or to synthesize in
pure form because of the resulting low yields. Thus, simple approaches for the
generation of such glucosides would be highly beneficial. We purified and
characterized a novel glucosyltransferase from plant cell suspension cultures of
Rauvolfia serpentina, which showed rather low substrate specificity. We obtained
its cDNA and expressed the active recombinant protein in bacteria (Escherichia
coli) with excellent plant/specific glucosylation efficiencies. Compared with
the plant system, the bacteria delivered the new enzyme, which was in the form
of a soluble or matrix/bound enzyme, approximately 1800 times more efficiently
for the synthesis of a wide range of glucosides. More importantly, the
engineered E. coli strain allowed for in vivo glucosylation and release of the
product into the culture medium, as shown by the formation of arbutin, which is
a potent inhibitor of human melanin biosynthesis with commercial value.
Copyright 2001 John Wiley & Sons, Inc.
Publication Types:
Comparative Study
Research Support, Non/U.S. Gov't
PMID: 11505382 [PubMed / indexed for MEDLINE]
26: J Nat Prod. 2001 May;64(5):686/7.
Revision of the structure of ajmalimine.
Hanhinen P, Lounasmaa M.
Laboratory for Organic and Bioorganic Chemistry, Technical University of
Helsinki, P.O. Box 6100, FIN/02015 HUT Espoo, Finland.
Careful reexamination of the published 1H and 13C NMR spectral data of
ajmalimine (1), an indole alkaloid from the roots of Rauvolfia serpentina,
indicates that, in reality, the compound is
(+)/17R/O/(3',4',5'/trimethoxybenzoyl)ajmaline (3).
PMID: 11374979 [PubMed / indexed for MEDLINE]
27: J Nat Prod. 2001 Jan;64(1):114/6.
3/oxo/rhazinilam: a new indole alkaloid from Rauvolfia serpentina x Rhazya
stricta hybrid plant cell cultures.
Gerasimenko I, Sheludko Y, Stockigt J.
Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes
Gutenberg/University, Staudinger Weg 5, D/55099 Mainz, Germany.
A new monoterpenoid indole alkaloid, 3/oxo/rhazinilam (1), was isolated from
intergeneric somatic hybrid cell cultures of Rauvolfia serpentina and Rhazya
stricta, and the structure was determined by detailed 1D and 2D NMR analysis. It
was also proved that 3/oxo/rhazinilam (1) is a natural constituent of the hybrid
cells.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 11170683 [PubMed / indexed for MEDLINE]
28: Planta Med. 2000 Oct;66(7):656/9.
Divergence of the indole alkaloid pattern in two somatic hybrid plant cell
subcultures of Rauvolfia serpentina x Rhazya stricta.
Sheludko Y, Gerasimenko I, Platonova O.
The alkaloid pattern in Rauvolfia serpentina x Rhazya stricta somatic hybrid
cell subcultures R x R17 K was studied and 11 compounds were identified on the
basis of their spectral data. Among them, 1,2/dehydroaspidospermidine,
rhazinilam, stemmadenine and tabersonine were reported as typical of Rhazya
species while vomilenine and sarpagine are characteristic of Rauvolfia alkaloid
metabolism. The alkaloid pattern in R x R17 K subcultures was compared with that
in the other hybrid cell subcultures (R x R17 M) which were developed from the
same origin hybrid cultures but have been maintained separately for about ten
years. The data presented here exhibit pronounced divergence of the alkaloid
patterns in R x R17 K and R x R17 M cell subcultures.
Publication Types:
Letter
Research Support, Non/U.S. Gov't
PMID: 11105575 [PubMed / indexed for MEDLINE]
29: Phytochemistry. 2000 Aug;54(7):657/66.
Molecular cloning and functional bacterial expression of a plant glucosidase
specifically involved in alkaloid biosynthesis.
Warzecha H, Gerasimenko I, Kutchan TM, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Institut fur Pharmazie, Johannes
Gutenberg/Universitat Mainz, Germany.
Monoterpenoid indole alkaloids are a vast and structurally complex group of
plant secondary compounds. In contrast to other groups of plant products which
produce many glycosides, indole alkaloids rarely occur as glucosides. Plants of
Rauvolfia serpentina accumulate ajmaline as a major alkaloid, whereas cell
suspension cultures of Rauvolfia mainly accumulate the glucoalkaloid
raucaffricine at levels of 1.6 g/l. Cell cultures do contain a specific
glucosidase. known as raucaffricine/O/beta/D/glucosidase (RG), which catalyzes
the in vitro formation of vomilenine, a direct intermediate in ajmaline
biosynthesis. Here, we describe the molecular cloning and functional expression
of this enzyme in Escherichia coli. RG shows up to 60% amino acid identity with
other glucosidases of plant origin and it shares several sequence motifs with
family 1 glucosidases which have been characterized. The best substrate
specificity for recombinant RG was raucaffricine (KM 1.3 mM, Vmax 0.5
nkat/microg protein) and only a few closely related structural derivatives were
also hydrolyzed. Moreover, an early intermediate of ajmaline biosynthesis,
strictosidine, is a substrate for recombinant RG (KM 1.8 mM, Vmax 2.6
pkat/microg protein) which was not observed for the low amounts of enzyme
isolated from Rauvolfia cells.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 10975500 [PubMed / indexed for MEDLINE]
30: Plant Physiol. 2000 Mar;122(3):793/801.
Detoxification of arsenic by phytochelatins in plants.
Schmoger ME, Oven M, Grill E.
Lehrstuhl fur Botanik, Technische Universitat Munchen, Biologikum/Weihenstephan,
Am Hochanger 4, 85350 Freising, Germany.
As is a ubiquitous element present in the atmosphere as well as in the aquatic
and terrestrial environments. Arsenite and arsenate are the major forms of As
intoxication, and these anions are readily taken up by plants. Both anions
efficiently induce the biosynthesis of phytochelatins (PCs)
([gamma/glutamate/cysteine](n)/glycine) in vivo and in vitro. The rapid
induction of the metal/binding PCs has been observed in cell suspension cultures
of Rauvolfia serpentina, in seedlings of Arabidopsis, and in enzyme preparations
of Silene vulgaris upon challenge to arsenicals. The rate of PC formation in
enzyme preparations was lower compared with Cd/induced biosynthesis, but was
accompanied by a prolonged induction phase that resulted finally in higher
peptide levels. An approximately 3:1 ratio of the sulfhydryl groups from PCs to
As is compatible with reported As/glutathione complexes. The identity of the
As/induced PCs and of reconstituted metal/peptide complexes has unequivocally
been demonstrated by electrospray ionization mass spectroscopy. Gel filtration
experiments and inhibitor studies also indicate a complexation and
detoxification of As by the induced PCs.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 10712543 [PubMed / indexed for MEDLINE]
31: Eur J Biochem. 2000 Mar;267(5):1397/406.
The gene encoding polyneuridine aldehyde esterase of monoterpenoid indole
alkaloid biosynthesis in plants is an ortholog of the alpha/betahydrolase super
family.
Dogru E, Warzecha H, Seibel F, Haebel S, Lottspeich F, Stockigt J.
Lehrstuhl fur Pharmazeutische Biologie, Institut fur Pharmazie, Johannes
Gutenberg/Universitat Mainz.
The biosynthesis of the anti/arrhythmic alkaloid ajmaline is catalysed by more
than 10 specific enzymes. In this multistep process polyneuridine aldehyde
esterase (PNAE) catalyses a central reaction by transforming polyneuridine
aldehyde into epi/vellosimine, which is the immediate precursor for the
synthesis of the ajmalane skeleton. PNAE was purified from cell suspension
cultures of Rauvolfia serpentina. The N/terminal sequence and endoproteinase
LysC fragments of the purified protein were used for primer design and for the
amplification of specific PCR products leading to the isolation of PNAE/encoding
cDNA from a R. serpentina library. The PNAE cDNA was fused with a C/terminal
His/tag, expressed in Escherichia coli and purified to homogeneity using
Ni/affinity chromatography. The pure enzyme shows extraordinary substrate
specificity, completely different to other esterases. Sequence alignments
indicate that PNAE is a new member of the alpha/beta hydrolase super family.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 10691977 [PubMed / indexed for MEDLINE]
32: Planta Med. 1996 Aug;62(4):358/9.
In vitro propagation of Rauvolfia serpentina through tissue culture.
Sarker KP, Islam A, Islam R, Hoque A, Joarder OI.
Multiple shoots were induced from nodal segments and shoot apices of Rauvolfia
serpentina and MS medium containing 1.0 mg/l BA and 0.1 mg/l NAA was found to
give the best shoot proliferation rate. Callus formed at cut bases of the
explants which produced shoots when subcultured on media containing low
concentration of BA (0.5 or 0.1 mg/l) and NAA (0.1 mg/l). The in vitro
proliferated shoots were rooted and later transferred to the soil.
Publication Types:
Letter
PMID: 8792671 [PubMed / indexed for MEDLINE]
33: Phytochemistry. 1994 Jan;35(2):353/60.
Heterologous expression of the plant proteins strictosidine synthase and
berberine bridge enzyme in insect cell culture.
Kutchan TM, Bock A, Dittrich H.
Laboratorium fur Molekulare Biologie, Universitat Munchen, Germany.
The heterologous expression of cDNAs encoding the alkaloid biosynthetic enzymes,
strictosidine synthase [EC 4.3.3.2] from Rauvolfia serpentina and the berberine
bridge enzyme [(S)/reticuline: oxygen oxidoreductase (methylene/bridge/forming),
EC 1.5.3.9] from Eschscholtzia californica, has been achieved in a cell culture
(Sf9) of the fall army worm, Spodoptera frugiperda, using a baculovirus/based
expression system. The expression resulted in the overproduction of each plant
enzyme in a catalytically active form. The maximal production attained was 4 mg
purified, active enzyme per litre cell culture for both the strictosidine
synthase and berberine bridge enzymes.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 7764480 [PubMed / indexed for MEDLINE]
34: Arch Biochem Biophys. 1992 May 1;294(2):717/23.
Strictosidine synthase from Rauvolfia serpentina: analysis of a gene involved in
indole alkaloid biosynthesis.
Bracher D, Kutchan TM.
Lehrstuhl fur Pharmazeutische Biologie, Universitat Munchen, Germany.
The gene for strictosidine synthase (str1), the enzyme which catalyzes the
stereospecific condensation of tryptamine and secologanin to form the key indole
alkaloid 3 alpha(S)/strictosidine has been isolated from genomic libraries
prepared from Rauvolfia serpentina (India) and from Rauvolfia mannii (West
Africa). The gene, str1, contained no introns and showed 100% nucleotide
sequence homology over 1180 bp, encompassing the entire reading frame, between
the two species. Transcription of the R. serpentina gene was found to start 81
nucleotides upstream from the AUG (26 nucleotides downstream from the TATA box).
Transient expression assays in Nicotiana plumbaginifolia protoplasts of the R.
serpentina str1 5'/noncoding region fused to the beta/glucuronidase reporter
gene revealed promoter activity equivalent to 4 +// 2% of that of 35 S CaMV
promoter control. A series of truncated segments of the str1 promoter region
indicated the presence of three areas of slight, but reproducible, negative
control. Gel retardation assays demonstrated that several regions of the
5'/flanking sequences specifically bound nuclear protein from R. serpentina and
that at least one region does not bind R. mannii nuclear protein. A survey of
the expression of str1 in the R. serpentina plant suggested that strictosidine
synthase poly(A)+ RNA was present predominantly, but not exclusively, in the
root. This result correlated well with the distribution of both enzyme activity
and indole alkaloids which were also predominant in the root, but, in general,
distributed throughout the shrub.
Publication Types:
Comparative Study
Research Support, Non/U.S. Gov't
PMID: 1567228 [PubMed / indexed for MEDLINE]
35: FEBS Lett. 1989 Oct 23;257(1):127/30.
Expression of enzymatically active cloned strictosidine synthase from the higher
plant Rauvolfia serpentina in Escherichia coli.
Kutchan TM.
Lehrstuhl fur Pharmazeutische Biologie, Universitat Munchen, FRG.
The cDNA for strictosidine synthase, the enzyme catalyzing the stereospecific
condensation of tryptamine with secologanin producing strictosidine, the key
intermediate in indole alkaloid biosynthesis, has been expressed in an
enzymatically active form in Escherichia coli. The cDNA trimmed of its 3'/ and
5'/flanking regions was inserted into the vector pKK223/3 by addition of a
synthetic adapter containing the ribosome binding site derived from the
beta/galactosidase gene. Strictosidine synthase activity (138 nkat.l/1) could be
measured in both whole bacteria and in bacterial protein extracts. Strictosidine
synthase represents the first enzyme of plant secondary metabolism to be
actively expressed in a microorganism.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 2680603 [PubMed / indexed for MEDLINE]
36: FEBS Lett. 1988 Sep 12;237(1/2):40/4.
The cDNA clone for strictosidine synthase from Rauvolfia serpentina. DNA
sequence determination and expression in Escherichia coli.
Kutchan TM, Hampp N, Lottspeich F, Beyreuther K, Zenk MH.
Lehrstuhl fur Pharmazeutische Biologie, Universitat Munchen, FRG.
The cDNA clone for strictosidine synthase, the enzyme which catalyzes the
stereospecific condensation of tryptamine with secologanin to form the key
intermediate in indole alkaloid biosynthesis, strictosidine, has been identified
with a synthetic oligodeoxynucleotide hybridization probe in a lambda gt11 cDNA
library of cultured cells of Rauvolfia serpentina. The DNA has been sequenced,
revealing an open reading frame of 1032 base pairs encoding 344 amino acids. The
sequence of 60 nucleotides in the 5'/flanking region has been determined by
primer extension analysis. The encoded protein has been expressed in E. coli DH5
as detected by immunoblotting of protein extracts with antibodies raised against
the native enzyme.
Publication Types:
Research Support, Non/U.S. Gov't
PMID: 3049153 [PubMed / indexed for MEDLINE]
37: Acta Pharmacol Toxicol (Copenh). 1986;59 Suppl 7:344/7.
Role of elements in pathophysiology of hypertension and antihypertensive drug
development.
Arora RB, Roy S, Khan SU.
Cadmium and Zinc levels were determined in 100 patients of Essential
hypertension by Atomic Absorption Spectrophotometer. It was shown that the mean
levels of Serum cadmium were 43.34% +// 6.5% higher and zinc were 28.42% +//
5.4% lower in hypertensive when compared with normotensive controls. A
statistically significant relationship between the height of diastolic blood
pressure and Serum cadmium levels was observed. Ajmaloon a preparation from
Rauvolfia serpentina with corrective herbs lowered the blood pressure
effectively and significantly P less than 0.001. The side effects were minimal.
It also tends to decrease the elevated serum cadmium levels in hypertensive
individuals. A plea for development of Catecholamine depleting agents as drug
for hypertension is advanced.
PMID: 3776586 [PubMed / indexed for MEDLINE]
38: Indian J Med Sci. 1957 Jul;11(7):479/82.
Effect of reserpine free extract of Rauvolfia serpentina on gastric acidity in
man.
RINDANI TH, ANTIA FP.
PMID: 13462576 [PubMed / OLDMEDLINE]
39: Arch Int Pharmacodyn Ther. 1956 Oct 1;108(1):51/4.
Influence of reserpine free extract of Rauvolfia serpentina on the action of
ACTH and cortisone acetate in rats.
RINDANI TH.
PMID: 13382379 [PubMed / OLDMEDLINE]
40: Wien Med Wochenschr. 1953 Jun 6;103(23):430/1.
[Rauvolfia serpentina in treatment of hypertension.]
[Article in Undetermined Language]
KLAUSGRABER F.
PMID: 13078578 [PubMed / indexed for MEDLINE]
