scholarly journals L-Amino Acid Ligase from Pseudomonas syringae Producing Tabtoxin Can Be Used for Enzymatic Synthesis of Various Functional Peptides

2013 ◽  
Vol 79 (16) ◽  
pp. 5023-5029 ◽  
Author(s):  
T. Arai ◽  
Y. Arimura ◽  
S. Ishikura ◽  
K. Kino
Keyword(s):  
Amino Acid ◽  
Pseudomonas Syringae ◽  
Enzymatic Synthesis ◽  
Functional Peptides

Characterization of an isozyme of S-adenosylmethionine synthetase from rat liver

1984 ◽  
Vol 62 (5) ◽  
pp. 276-279 ◽  
Author(s):  
C. H. Lin ◽  
W. Chung ◽  
K. P. Strickland ◽  
A. J. Hudson
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Amino Acid ◽  
Enzymatic Synthesis ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Aromatic Amino Acids ◽  
Adenosylmethionine Synthetase ◽  
Cellulose Column

An isozyme of S-adenosylmethionine synthetase has been purified to homogeneity by ammonium sulfate fractionation, DEAE-cellulose column chromatography, and gel filtration on a Sephadex G-200 column. The purified enzyme is very unstable and has a molecular weight of 120 000 consisting of two identical subunits. Amino acid analysis on the purified enzyme showed glycine, glutamate, and aspartate to be the most abundant and the aromatic amino acids to be the least abundant. It possesses tripolyphosphatase activity which can be stimulated five to six times by S-adenosylmethionine (20–40 μM). The findings support the conclusion that an enzyme-bound tripolyphosphate is an obligatory intermediate in the enzymatic synthesis of S-adenosylmethionine from ATP and methionine.

scholarly journals Identification of the Biosynthetic Gene Cluster for 3-Methylarginine, a Toxin Produced by Pseudomonas syringae pv. syringae 22d/93

2010 ◽  
Vol 76 (8) ◽  
pp. 2500-2508 ◽  
Author(s):  
S. D. Braun ◽  
J. Hofmann ◽  
A. Wensing ◽  
M. S. Ullrich ◽  
H. Weingart ◽  
...  
Keyword(s):  
Amino Acid ◽  
Gene Cluster ◽  
Pseudomonas Syringae ◽  
Biosynthetic Gene Cluster ◽  
Pentanoic Acid ◽  
Promising Candidate ◽  
Biosynthesis Gene Cluster ◽  
E Coli ◽  
Highly Active ◽  
Putative Aminotransferase

ABSTRACT The epiphyte Pseudomonas syringae pv. syringae 22d/93 (Pss22d) produces the rare amino acid 3-methylarginine (MeArg), which is highly active against the closely related soybean pathogen Pseudomonas syringae pv. glycinea. Since these pathogens compete for the same habitat, Pss22d is a promising candidate for biocontrol of P. syringae pv. glycinea. The MeArg biosynthesis gene cluster codes for the S-adenosylmethionine (SAM)-dependent methyltransferase MrsA, the putative aminotransferase MrsB, and the amino acid exporter MrsC. Transfer of the whole gene cluster into Escherichia coli resulted in heterologous production of MeArg. The methyltransferase MrsA was overexpressed in E. coli as a His-tagged protein and functionally characterized (Km , 7 mM; k cat, 85 min−1). The highly selective methyltransferase MrsA transfers the methyl group from SAM into 5-guanidino-2-oxo-pentanoic acid to yield 5-guanidino-3-methyl-2-oxo-pentanoic acid, which then only needs to be transaminated to result in the antibiotic MeArg.

scholarly journals Direct Enzymatic Synthesis of a Deep‐Blue Fluorescent Noncanonical Amino Acid from Azulene and Serine

ChemBioChem ◽  
2019 ◽  
Vol 21 (1-2) ◽  
pp. 80-83 ◽  
Author(s):  
Ella J. Watkins ◽  
Patrick J. Almhjell ◽  
Frances H. Arnold
Keyword(s):  
Amino Acid ◽  
Enzymatic Synthesis ◽  
Deep Blue ◽  
Noncanonical Amino Acid

Enzymatic synthesis of Z-aspartame in liquefied amino acid substrates

2013 ◽  
Vol 70 ◽  
pp. 84-87 ◽  
Author(s):  
Shinya Furukawa ◽  
Kazuhiro Hasegawa ◽  
Ichiro Fuke ◽  
Koji Kittaka ◽  
Terumitsu Nakakoba ◽  
...  
Keyword(s):  
Amino Acid ◽  
Enzymatic Synthesis

Investigations of the biosynthesis of the phytotoxin coronatine

1994 ◽  
Vol 72 (1) ◽  
pp. 86-99 ◽  
Author(s):  
Ronald J. Parry ◽  
Sunil V. Mhaskar ◽  
Ming-Teh Lin ◽  
Alan E. Walker ◽  
Robson Mafoti
Keyword(s):  
Amino Acid ◽  
Pseudomonas Syringae ◽  
Cyclopropane Ring ◽  
Carbonyl Oxygen ◽  
Oxidative Cyclization ◽  
Amide Bond ◽  
Hydrogen Atoms ◽  
Coronafacic Acid ◽  
Polyketide Chain ◽  
Oxygen Atoms

The biosynthesis of the phytotoxin coronatine has been investigated by administration of isotopically labeled precursors to Pseudomonas syringae pv. glycinea. The structure of coronatine contains two moieties of distinct biosynthetic origin, a bicyclic, hydrindanone carboxylic acid (coronafacic acid) and a cyclopropyl α-amino acid (coronamic acid). Investigations of coronafacic acid biosynthesis have shown that this compound is a polyketide derived from three acetate units, one butyrate unit, and one pyruvate unit. The two carbonyl oxygen atoms of coronafacic acid were found to be derived from the oxygen atoms of acetate. Additional experiments are described that rule out some possible modes for assembly of the polyketide chain. Coronamic acid is shown to be derived from L-isoleucine via the intermediacy of L-alloisoleucine. Examination of the mechanism of the cyclization of L-alloisoleucine to coronamic acid revealed that the formation of the cyclopropane ring takes place with the removal of only two hydrogen atoms from the amino acid, one at C-2 and the other at C-6. The nitrogen atom at C-2 of L-alloisoleucine is shown to be retained. On the basis of these observations, a mechanism is postulated for the cyclization reaction that involves the diversion of an enzymatic hydroxylation reaction into an oxidative cyclization. Finally, a precursor incorporation experiment with deuterium-labeled coronamic acid demonstrated that free coronamic acid can be efficiently incorporated into coronatine. This observation indicates that the cyclization of L-alloisoleucine to coronamic acid can occur before formation of the amide bond between coronafacic acid and coronamic acid.

scholarly journals Sequence Diversity of rulA among Natural Isolates of Pseudomonas syringae and Effect on Function of rulAB-Mediated UV Radiation Tolerance

2000 ◽  
Vol 66 (12) ◽  
pp. 5167-5173 ◽  
Author(s):  
George W. Sundin ◽  
Janette L. Jacobs ◽  
Jes�s Murillo
Keyword(s):  
Amino Acid ◽  
Pseudomonas Syringae ◽  
Uv Radiation ◽  
Radiation Sensitivity ◽  
Amino Acid Sequences ◽  
Radiation Tolerance ◽  
Plant Host ◽  
Long Time ◽  
Natural Isolates ◽  
Wavelength Radiation

ABSTRACT The rulAB locus confers tolerance to UV radiation and is borne on plasmids of the pPT23A family in Pseudomonas syringae. We sequenced 14 rulA alleles from P. syringae strains representing seven pathovars and found sequence differences of 1 to 12% within pathovar syringae, and up to 15% differences between pathovars. Since the sequence variation withinrulA was similar to that of P. syringaechromosomal alleles, we hypothesized that rulAB has evolved over a long time period in P. syringae. A phylogenetic analysis of the deduced amino acid sequences of rulAresulted in seven clusters. Strains from the same plant host grouped together in three cases; however, strains from different pathovars grouped together in two cases. In particular, the rulAalleles from P. syringae pv. lachrymans and P. syringae pv. pisi were grouped but were clearly distinct from the other sequenced alleles, suggesting the possibility of a recent interpathovar transfer. We constructed chimeric rulABexpression clones and found that the observed sequence differences resulted in significant differences in UV (wavelength) radiation sensitivity. Our results suggest that specific amino acid changes in RulA could alter UV radiation tolerance and the competitiveness of theP. syringae host in the phyllosphere.

Enzymatic synthesis of amino acid ester of butyl α-D-glucopyranoside

1994 ◽  
Vol 35 (21) ◽  
pp. 3535-3536 ◽  
Author(s):  
Jean Fabre ◽  
François Paul ◽  
Pierre Monsan ◽  
Casimir Blonski ◽  
Jacques Périé
Keyword(s):  
Amino Acid ◽  
Enzymatic Synthesis ◽  
Acid Ester ◽  
Amino Acid Ester

Enzymatic synthesis of the neuroexcitatory amino acid quisqualic acid by cysteine synthase

1986 ◽  
Vol 25 (12) ◽  
pp. 2759-2763 ◽  
Author(s):  
Isamu Murakoshi ◽  
Masakazu Kaneko ◽  
Chiharu Koide ◽  
Fumio Ikegami
Keyword(s):  
Amino Acid ◽  
Enzymatic Synthesis ◽  
Cysteine Synthase ◽  
Quisqualic Acid

scholarly journals Characterisation, Manipulation and Directed Evolution of Non-Ribosomal Peptide Synthetase Enzymes

2021 ◽  
Author(s):  
◽  
Jeremy George Owen
Keyword(s):  
Amino Acid ◽  
Directed Evolution ◽  
Pseudomonas Syringae ◽  
Gene Knockout ◽  
Iron Starvation ◽  
Biochemical Characterisation ◽  
T Domain ◽  
Stereo Selectivity

<p><b>Non-ribosomal peptide synthetases (NRPS) are large, modular enzymes that synthesisebiologically active secondary metabolites from amino acid precursors without the need for anucleic acid template. NRPS play an integral role in microbial physiology and also havepotential applications in the synthesis of novel peptide molecules. Both of these aspects areexamined in this thesis.</b></p> <p>Under conditions of iron starvation Pseudomonas syringae synthesises siderophores for activeuptake of iron. The primary siderophore of P. syringae is pyoverdine, a fluorescent moleculethat is assembled from amino acid (aa) precursors by NRPS. Five putative pyoverdine NRPSgenes in P. syringae pv. phaseolicola 1448a (Ps1448a) were identified and characterised insilico and their role in pyoverdine biosynthesis was confirmed by gene knockout. Creation ofpyoverdine null Ps1448a enabled identification of a previously uncharacterised temperatureregulatedsecondary siderophore, achromobactin, which is NRPS independent and has loweraffinity for iron. Pyoverdine and achromobactin null mutants were characterised in regard toiron uptake, virulence and growth in iron-limited conditions. Determination of the substratespecificity for the seven adenylation (A) domains of the Ps1448a pyoverdine sidechain NRPSwas also attempted. Although ultimately unsuccessful, these attempts provided a rigorousassessment of methods for the expression, purification and biochemical characterisation of Adomains.</p> <p>The Ps1448a NRPS were subsequently employed in domain swapping experiments to testcondensation (C) domain specificity for aa substrates during peptide formation in vivo.</p> <p>Experiments in which the terminal C- and/or A-domain of the Pseudomonas aeruginosa(PAO1) pyoverdine NRPS system were replaced with alternative domains from Ps1448a andPAO1 were consistent with previous in vitro observations that C-domains exhibit strongsidechain and stereo-selectivity at the downstream aa position, but only stereo-selectivity atthe upstream aa position.</p> <p>These results prompted investigation into the role of inter-domain communication in NRPSfunction, to test the hypothesis that the thiolation (T) domain enters into specific interactionswith other domains, which might provide an alternative explanation for the diminished activityof recombinant NRPS enzymes. A recently characterised single-module NRPS, bpsA, waschosen as a reporter gene for these experiments based on its ability to generate blue pigment inEscherichia coli. Substitution of the native bpsA T-domain consistently impaired function,consistent with the hypothesis. It was shown that directed evolution could be applied to restorefunction in substituted T-domains. Mutations that restored function were mapped in silico, anda structural model for interaction between the thioester (TE) and T-domain of BpsA wasderived.</p> <p>The utility of bpsA for discovery and characterisation of phosphopantetheinyl transferase(PPTase) enzymes was also investigated. In vivo and in vitro assays for determination ofPPTase activity were developed and a high-throughput screen for discovery of new PPTases inenvironmental DNA libraries was successfully implemented.</p>

Enzymatic Synthesis of Peptides with Hydrophilic Amino Acid in Water-Miscible Organic Solvent

1992 ◽  
Vol 672 (1 Enzyme Engine) ◽  
pp. 458-461 ◽  
Author(s):  
YUKTTAKA KIMURA ◽  
YOSHIKO TARI ◽  
SHUJI ADACHI ◽  
RYUICHI MATSUNO
Keyword(s):  
Amino Acid ◽  
Organic Solvent ◽  
Enzymatic Synthesis ◽  
Hydrophilic Amino Acid
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