Clostridium sticklandii

Arthrobacter hjalinus Pacillus megaterium Butyribacterium rettgeri Clostridium sticklandii Clostridium tetanomorphum Clostridium thermoaceticum... 

Glycine reductase selenoprotein A Energy metabolism Clostridium sticklandii 5, 99, 100... 

D-proline reductase Anaerobic respiration Clostridium sticklandii 104, 105... 

Clostridium sticklandii also expresses a proline reductase that can reduc-tively cleave proline to 8-aminovalerate (Seto and Stadtman 1976). PR was first purified by Seto and Stadtman (1976) by following the decomposition of proUne in the presence of dithiothreitol or NADH. They found PR to have a denatured mass of approximately 30kDa (sodium dodecyl sulfate-polyacrylomide gel electrophoresis SDS-PAGE) and a native size of approximately 300 kDa. The addition of selenite to the growth medium of C sticklandii did increase the specific activity of PR in extracts by threefold however, no selenium was detected in the purified enzyme. It should be noted that this purified enzyme had lost the ability to couple reduction of proline to NADH and thus probably was missing one or more components of the complete enzyme complex. 

Seto B. 1980. Chemical characterization of an alkali-labile bond in the polypeptide of proline reductase from Clostridium sticklandii. J Biol Chem 255 5004-6. 

Sliwkowski MX, StadtmanTC. 1988. Selenium-dependent glycine reductase differences in physicochemical properties and biological activities of selenoprotein A components isolated from Clostridium sticklandii and Clostridium purinolyticum. Biofactors 1 293-6. 

Stadtman TC, Elliott P. 1957. Studies on the Enzymatic Reduction of amino acids II. Purification and properties of ao-proline reductase and a prohne racemase from Clostridium sticklandii. 1 Biol Chem 228 983-97. 

Scheme 1.6.16. Biosynthesis of (3S,5S)-diaminohexanoic acid in Clostridium sticklandii.

Besides a lysine 5,6-aminomutase, Clostridium sticklandii also has a D-ornithine 4,5-aminomutase (EC 5.4.3.5) [78, 79], D-Ornithine is generated from L-ornithine by ornithine racemase [80], The two genes encoding D-ornithine 4,5-aminomutase have been cloned, sequenced, and expressed in E. coli [81]. The enzyme is an a2/ 2-heterotetramer, consisting of 12 800 Da and 82 900 Da subunits. The protein requires Bn and pyridoxal phosphate as cofactors. Similar to the lysine 5,6-aminomutase, a conserved base-ofi)/histidine-on cobalamin binding motif is present in the 82 900 Da protein. 

Lysine mutase is the first of a group of AdoCbl-dependent enzymes that catalyses the 1,2-migration of an amino group (Fig. 26). It has been isolated from Clostridium sticklandii [39] and consists of a cobalamin-binding orange protein and a smaller yellow protein. Apart from AdoCbl, several other essential cofactors have been identified, such as pyridoxal phosphate, ATP, FAD, thiols, Mg2+ and K+ [38]. The function of the yellow protein and some of these cofactors is to renew continuously... 

Other pyruvate-containing enzymes include aspartate /3-decarboxylase from Escherichia coli, the enzyme that cattdyzes the formation of /3-alanine for the synthesis of pantothenic acid (Section 12.2.4) proline reductase from Clostridium sticklandii phosphatidylserine decarboxylase from E. coli and phenyltdtuiine tuninoticmsferase from Pseudomonas fluorescens. Phospho-pemtetheinoyl cysteine deceu boxylase, involved in the synthesis of coenzyme A (Section 12.2.1), and S-adenosylmethionine decarboxylase seem to be the only mammtditm pyruvoyl enzymes (SneU, 1990). 

Proline racemase is a member of a broad family of cofactor-independent epimerases and racemases, and has been very well characterized mechanistically. The proline racemase from Clostridium sticklandii was the first of the cofactor-independent racemases to be characterized [13, 80], The enzyme participates in the catabolism of L-proline, producing o-proline as a substrate for o-proline oxidase [4]. Early... 

Chang, C. H., Frey, P. A. (2000) Cloning, sequencing, heterologous expression, purification, and characterization of adenosylcobalamin-dependent D-lysine 5,6-aminomutase from Clostridium sticklandii, J. Biol. Chem. 275, 106-114. 

The Stickland reaction (47) has received much attention as a possible route to chiral acetate due to the availability of chiral glycine (48). In the Stickland reaction two moles of glycine and one mole of d-alanine are converted quantitatively into three moles of acetate, three moles of ammonia, and one mole of C02 by the organism Clostridium sticklandii. The presence of amino acid transaminase in the intact organisms leads to extensive hydrogen exchange although in the purified enzyme the replacement of NH2 by H occurs stereospecifically with inversion (49, 50). Unfortunately, the rates of conversion with the purified enzyme are too low to be synthetically useful. 

Proline racemase occurs in Clostridium sticklandii, which produces 8-aminovalerate from L-proline. Proline racemase and D-proline reductase are responsible for the conversion L-proline is racemized by proline racemase to form D-proline, which is converted into 5-aminovalerate by D-proline reductase (E.C. 1.4.4.1). 

Rudnick and Abeles purified proline racemase to 95% homogeneity from Clostridium sticklandii, and characterized it 92. The enzyme is composed of two identical subunits with a molecular weight of about 38000, and is independent of any cofactors or metals. Most amino acid racemases require pyridoxal 5 -phosphate, which labilizes the bond between the a-hydrogen and the chiral center by aldimine formation with the a-amino group of the substrate. However, PLP is not involved in the reaction of proline racemase acting on an a-imino acid. The enzyme also acts on 2-hydroxy-L-proline and 2-allo-hydroxy-D-proline although slowly they are epimer-ized at a rate of 2 and 5% of the rate of L-proline racemization, respectively. L-Proline and D-proline showed Km values of 2.9 and 2.5 mti, respectively1119. ... 

Yagasaki and Ozaki 126 developed a method for production of D-proline from l-proline using the recombinant proline racemase of Clostridium sticklandii. L-Proline was degraded by Candida sp. PRD-234, and optically pure D-proline was obtained. 

Use of serine hydroxymethyltransferase with [2,2- H2]glycine and or [2,2- H2]glycine and H20 gives (2S)- and (2/ )-[2- H 2- Hj]glycines, respectively, and these were incubated with glycine reductase from Clostridium sticklandii to yield chirally labeled acetates (62) (Scheme 14). These acetates were assayed as described in references 3 and 7 to show that the reduction proceeded with inversion of configuration (62). 

A preparation (Wright and Anderson, 1958) from Clostridium sticklandii catalyzed the reaction PGA + pyruvate + CoA — acetyl CoA + FH2. Since the diglutamyl and triglutamyl pterins and their J T -formyl derivatives were reduced, such a reductase system can account for the transformation of exogenous PGA into the reduced coenzymatic products. An enzyme from Micrococcus lactilyticus formed FH2 the reaction required an electron donor such as pyruvate or a-ketobutyrate, a dithiol such as dimer-captopropanol, orthophosphate, and reduced CoA. This enzyme did not form FH4 derivatives. 

The effectiveness of phytofactors influencing bacteria specialized in ammonia production depends also on their chemical structure, which was proved by Flythe and Kagan (39). They investigated red clover Triforium pratense), rich in soluble phenols, and for direct experiments they used extracted pure phenolic compounds, such as biochanin A, and also the extract from the whole plant The anti-effect of Clostridium sticklandii, one of the bacteria from the HAP group, was observed in the case of the extract from the whole plant and biochanin A, while no effect was found for other phenolic compounds. The results of research indicate that some... 

Paster, B.J., J.B. Russell, C.M.J. Yang, J.M. Chow, C.R. Woese and R. Tanner, 1993. Phylogeny of the ammonia-producing ruminal bacterial Peptostreptococcus anaerobius, Clostridium sticklandii, and Clostridium aminophilum spp. nov. International J. Systematic Bacteriol. 43, 107-110. 

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