Vitamin B6 Enzyme Research Articles

Harnessing the Value of Rumen Protected Amino Acids to Enhance Animal Performance – A Review

Abstract In general, higher mammals need nine amino acids (AA) in their diets as building blocks to synthesize proteins while ruminants can produce some of them through the synthesis of microbial proteins. Diet is utilized by ruminal microorganisms to synthesize microbial protein (MCP) which is digested in the small intestine (SI). Although protein and amino acid requirements in ruminants are subject to microbial protein synthesis, it is not enough for optimal daily production. Therefore, there is a current trend towards supplementing amino acids in ruminant diets. In the rumen, free amino acids can be degraded by rumen bacteria, therefore, the AAs need to be supplemented in a protected form to be stable in the rumen and absorbable post-ruminal for metabolic purposes. The main site of amino acid absorption is the small intestine (SI), and there is a need to keep AA from ruminal degradation and direct them to absorption sites. Several approaches have been suggested by feed scientists to decrease this problem such as defaunation and debacterization of the rumen against amino acid-fermenting fungi and bacteria, inhibitors or antagonists of vitamin B6 enzymes, diet composition and also protecting AA from rumen degradation. A number of studies have evaluated the roles of amino acids concerning their effects on milk yield, growth, digestibility, feed intake and efficiency of nitrogen utilization of ruminants. The focus of this review was on experimental and research studies about AAs in feedstuff, metabolism, supplementing amino acids for ruminants and the current trends of using rumen protected amino acids.

Synthesis, characterization, computational study, and biological relevance of a family of isostructural, mononuclear Ln (Ln = Gd, Tb, Dy, Ho, Er) complexes containing pyridoxine, an essential ingredient of vitamin B6 enzyme

The synthesis and characterization of a family of LnIII complexes (Ln=Gd(1), Tb(2), Dy(3), Ho(4) and Er(5)) of formula [(LnIII(NO3)2(OH2)(pyridoxine)2](NO3) are reported, where pyridoxine is 2-methyl-3-hydroxy-4,5-bis(hydroxymethyl)pyridine. They were obtained from the reaction of Ln(NO3)3 with pyridoxine in ethanol in a 1:1M ratio. The crystal structures of representative 1 and 5 were obtained using a single-crystal X-ray diffractometer. The subsequent characterization of all the complexes was performed using elemental analysis, IR spectroscopy and UV–Vis spectroscopy. The gadolinium complex was further analyzed to determine if DNA is a biological target of this series of metal complexes. In this regard, the interaction between the gadolinium complex and DNA was probed utilizing circular dichroism spectroscopy. The study supports the moderate binding of complex 1 to DNA.

Urine 3-hydroxykynurenine is higher during the postovulatory phase than in the preovulatory phase indicating a higher vitamin B6 requirement.

The relationship between l-tryptophan to nicotinamide metabolism and the menstrual cycle of Japanese women was investigated. Nine metabolism intermediates from urine samples collected during the preovulatory and postovulatory phases were measured. Only urine 3-hydroxykynurenine was higher in the postovulatory phase than in the preovulatory phase. This increase in 3-hydroxykynurenine suggests a decreased reaction of 3-hydroxykynurenine → 3-hydroxyanthranilic acid catalyzed by kynureninase, a vitamin B6 enzyme.

Vitamin B6 enzymes participating in selenium amino acid metabolism.

Various vitamin B6 enzymes play important roles in mammalian and microbial metabolism of selenium amino acids. Selenocysteine is synthesized from selenohomocysteine by catalysis of cystathionine beta-synthase and cystathionine gamma-lyase, which both require pyridoxal phosphate. Selenocysteine beta-lyase, a new B6-enzyme, exclusively catalyzes beta-elimination of selenocysteine, and occurs in mammalian systems and bacteria. Methionine gamma-lyase, cysteine desulfurase, cysteine sulfinate desulfinase, and D-selenocystine alpha,beta-lyase, which are B6-enzymes, act on cysteine, cysteine sulfinate, D-cystine, and their derivatives, and their selenium counterparts indiscriminately. Their reaction mechanisms are comparatively described.

Crystallization and preliminary X-ray analysis of the β-isoform of glutamate decarboxylase from Escherichia coli

Glutamate decarboxylase (GAD) is a vitamin B6 enzyme which catalyzes the alpha-decarboxylation of L-glutamate to gamma-aminobutyric acid (GABA). Escherichia coli cells coexpress two highly homologous enzyme isoforms, GADalpha and GADbeta. Well diffracting crystals of GADbeta were obtained by taking advantage of the possibility of expressing each isoform separately. They belong to space group P31 or P32 with the unit-cell dimensions a = b = 115.6 and c = 206.6 A and contain one GAD hexamer in the asymmetric unit. High-resolution synchrotron data were collected at 100 K for the native protein and a potential heavy-atom derivative.

Vitamin B‐6 model reactions. Part VI‐a Hammett relationship for the magnitude of pseudoallylic proton‐proton spin coupling constants in substituted azomethines

AbstractFour‐bond proton‐proton spin coupling constants were measured between the azomethine proton and the glycine CH protons of some coordinated glycine Schiff bases where the parent aldehyde is aromatic. Such systems are models for vitamin B6 enzyme substrate complexes. It is demonstrated that the coupling constants are not only related to the dihedral angles between coupled systems but they are also linearly related to the Hammett σ value of the substituents on the aromatic ring. A method is suggested for the calculation of dihedral angles between proton‐carbon bonds in these pseudoallylic systems.

β-Replacement Reaction in an Asymmetric Site Provided by an Artificial Enzyme Composed of a Hydrophobic Vitamin B6 and Peptide Lipids

Abstract An artificial vitamin B6 enzyme composed of a hydrophobic vitamin B6, a peptide lipid, and an additional peptide lipid having (S)-binaphtol and (S)-alanine moieties exhibited a strong CD band in a ultraviolet region in an aqueous medium, and mediated enantioselective formation of tryptophan in the presence of bivalent copper ions as a tryptophan synthase-like reaction.

Disruption of active site interactions with pyridoxal 5'-phosphate and substrates by conservative replacements in the glycine-rich loop of Escherichia coli D-serine dehydratase

We have used site-directed mutagenesis to examine the function of three putative active site residues (C278, G279, and G281) of the vitamin B6 enzyme D-serine dehydratase. These residues lie in or adjacent to a conserved glycine-rich loop that is known to interact with the pyridoxal 5'-phosphate cofactor in several B6 enzymes and that resembles the GXGXXG loop of nucleotide-binding sites. The cofactor affinity, catalytic properties, and spectral properties (UV, CD, fluorescence, and 31P NMR) of alanine variants C278A, G279A, and G281A were measured as well as the susceptibility of each variant to thiol modification by 5,5'-dithiobis(2-nitrobenzoic acid). The specific thiols modified in each variant and wild type D-serine dehydratase were identified by amino acid sequencing of labeled tryptic peptides. C278A, G279A, and G281A displayed 10-, 33-, and 22-fold lower affinities for pyridoxal 5'-phosphate than did wild type D-serine dehydratase and turnover numbers with D-serine that were 50, 6, and 60% of normal, respectively. The introduction of a methyl side chain into G281 enhanced catalytic efficiency with the substrates D-threonine, D-allo-threonine, and L-serine, whereas the methyl side chain at position 279 impaired catalysis of all substrates as well as cofactor affinity. The 31P NMR spectrum of D-serine dehydratase was minimally perturbed by the alanine substitutions, consistent with the view that neither G279 nor G281 interacts with the phosphate group of the cofactor (in contrast to the arrangement found in several other B6 enzymes). C311 was the single thiol modified by 5,5'-dithiobis(2-nitrobenzoic acid) in wild type D-serine dehydratase. Two normally inaccessible thiol groups, C233 and C278, were rendered susceptible to modification as a consequence of either G----A substitution, and modification of C278 was associated with inactivation of G279A and G281A. These observations suggest that small perturbations in the glycine-rich loop induce conformational changes spanning a considerable area around the active site.

Models of vitamin B6 enzymes. 3. Steric and electronic effects in carbon-hydrogen bond breaking reactions of bis(salicylideneglycinato)cobaltate(III) anions

Salicylaldehydes readily Schiff bases with glycine. These schiff bases form bis(salicylideneglycinato) cobaltate (III) complexes. Twelve cobalt (III) complexes have been synthesized with substituents variously on the 3-, 4-, 5-, and 6-positions of the aromatic ring of the salicylaldehyde moiety. The carbon-hydrogen bond breaking reactions of the two gem-methylenic protons of the glycine moiety have been studied by deuterium exchange. Rates are first order in complex and first order in hydroxide ion. Rates differ for the exchange of the two protons, and their ratio varies from 0.81 to 4.7. This demonstrates a reversal of stereoselectivity, depending on ring substituents. Rates vs substituents generally follow Hammett behavior. The structural and electronic features that lead to stereoselectivity are discussed

ニジマスの卵発生過程における水溶性ビタミン含有量およびトランスアミナーゼ活性の変化

To know a physiological role of vitamin in the development of fish eggs, changes in the con-tents of ascorbic acid, pantothenic acid, biotin, niacin and vitamin B6, and activities of glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) in developing rainbow trout eggs were determined. The ascorbic acid content rapidly decreased by half level on third day after fertilization, thereafter the vitamin tended to decrease during development. The sac fry, when a substantial amount of the yolk had disappeared, retained the ascorbic acid and was about one fourth of that of the initial eggs. The amounts of thiamine, pantothenic acid and biotin were reduced by half during the course of development, while niacin content tended to increase after hatching. The amounts of vitamin B6 decreased by about 45% during the course of the development. Some variation of the composition were found: the concentration of pyridoxal decreased appreciably, while those of pyridoxal 5'-phosphate, pyridoxamine 5'-phosphate and pyridoxamine increased in the course of development. The activities of GOT and GPT were very low in the early stages of development. After hatching, the activities of the vitamin B6 enzymes increased greatly.

The effects of administration of semicarbazide and aminooxyacetic acid on B6 vitamer levels in subcellular fractions of mouse brain.

The in vivo effects of semicarbazide (SC) and aminooxyacetic acid (AOAA) on the B6 vitamer concentrations in brain subcellular fractions, especially in extrasynaptosomal, synaptosomal plasma, extrasynaptosomal and synaptosomal mitochondria, were investigated. Treatment of mice with SC decreased levels of pyridoxal phosphate (PLP) in extrasynaptosomal and synaptosomal plasma, while it had no effect on those in extrasynaptosomal and synaptosomal mitochondria. On the contrary, treatment with AOAA more significantly decreased pyridoxamine phosphate (PMP) levels in extrasynaptosomal and synaptosomal mitochondria than those in extrasynaptosomal and synaptosomal plasma. According to these results, the difference between in vivo effects of SC and AOAA on vitamin B6-enzymes was discussed.

Coenzymatic activities of 2-nor-2-hydroxymethyl pyridoxal 5'-phosphate for various vitamin B6 enzymes.

The coenzymatic activities of 2-nor-2-hydroxymethyl-pyridoxal 5'-phosphate (2'-hydroxy PLP) for various vitamin B6-dependent enzymes were studied in order to obtain information on the role of the methyl group at position 2 of pyridoxal 5'-phosphate (PLP) for the binding to apoprotein and for the catalytic action of the holoenzyme. Although it has been demonstrated that this coenzyme analog binds to the catalytic center and serves as a coenzyme for a variety of B6 en-zymes, the mode of binding as well as the catalytic activity of the holo-enzyme reconstituted with the analog was different according to the kinds of enzyme and assay conditions. In the ordinary assay system using Tris-HCl buffer, 2'-hydroxy PLP showed almost an equal affinity for pig heart muscle mitochondrial and cytoplasmic aspartate aminotransferase (GOTs, and GOTm) to that of PLP. The maximal velocity (Vmax value) of the reaction mediated by GOTs, or GOTm reconstituted with 2'-hydroxy PLP was somewhat large as compared with that catalyzed by the native holoenzyme. How-ever, when phosphate buffer was used for the assay system, the affinity of the coenzyme analog decreased markedly and the Vmax value catalyzed by the analog-bound GOTs, or GOTm became approximately 70% of that by the native enzyme. Thus as judged from the Vmax value under ordinary assay conditions, 2'-hydroxy PLP is considered to be rather superior to PLP as the coenzyme for GOT. Phosphate anion inhibited the binding of PLP or 2'-hydroxy PLP to apoGOT in a competitive manner and further caused an appreciable decrease in the apparent Vmax of the reaction catalyzed by the resulting holoenzyme. The K1, values of phosphate anion against PLP and 2'-hydroxy PLP were 11.9 and 3.75, respectively. On the other hand, in the case of E. coli tryptophanase, both the affinity for 2'-hydroxy PLP and the Vmax value of the reaction mediated by the analog-bound enzyme were significantly smaller than those of the native coenzyme irrespective of the buffer used. Of the other B6 enzymes tested hitherto, E. intermedia tyrosine phenol-lyase and Pseudomonas dacunhae aspartate β-decarboxylase showed the same behaviors toward 2'-hydroxy PLP as that of tryptophanase. On the contrary, the coenzyme analog served as a more efficient co-enzyme than the native coenzyme for E. coli D-serine dehydrase in the presence of phosphate anion. These results indicate that there are at least three groups of B6 enzymes exhibiting different interaction with the 2-methyl group of PLP.

The reaction of amines with pyridoxal azomethines. The question of transaldimation and its role in the mechanism of vitamin B6 enzymes.

The reaction of amines with pyridoxal azomethines. The question of transaldimation and its role in the mechanism of vitamin B6 enzymes.

Tryptophan metabolism in patients with pellagra: problem of vitamin B6 enzyme activity and feedback control of tryptophan pyrrolase enzyme

Tryptophan metabolism in patients with pellagra: problem of vitamin B6 enzyme activity and feedback control of tryptophan pyrrolase enzyme

Inhibitory effect of beta-PIPH on some enzymatic activities related to vitamin B6

Inhibition of some vitamin B6 enzymes by carbonyl reagents, metal-chelating agents and phenylalanine derivatives has been reported. Recently, monamine oxidase inhibition by β-phenyl-isopropyl-hydrazine has also been demonstrated, but it is not clear whether monamine oxidase is related to vitamin B6 enzyme and vitamin B2 enzyme or not. In the present paper, we report on our studies, on the mechanism of inhibition of the tryptophanase (vitamin B6 enzyme) and d-amino acid oxidase (vitamin B2 enzyme) by β-phenyl-isopropyl-hydrazine.The results were as follows ;1) β-PIPH was found to be inhibitory on the tryptophanase, although it did not influence the inhibitory response on the d-amino acid oxidase.2) Detailed study indicated that β-PIPH is a nonspecific, formally noncompetitive inhibitor of the tryptophanase.3) This inhibition is achieved by the removal of cotryptophanase added to the system in vitro through nonenzymatic reaction with β-PIPH.4) This conclusion was strengthened by the preparation and characterization of the product of the reaction between pyridoxal and β-PIPH, and its identification as a substituted pyridoxylidene-β-phenyl-isopropyl-hydrazine.

Failure of berberine to inhibit pig heart glutamic-aspartic transaminase.

Previous studies from this laboratory have established that the alkaloid berberine inhibits two typical vitamin B6-enzymes of bacterial origins, i. e. tyrosine decarboxylase of Streptococcus faecalis and tryptophanase of Escherichia coli. Evidence has also been reported to indicate that berberine inhibits these enzymes by competing with the coenzyme pyridoxal phosphate. It has further been shown that the alkaloid, once combined with the apoenzymes, causes slow and irreversible inactivation of the enzymes.The present investigation was undertaken to examine if berberine behaves similarly toward pig heart glutamic-aspartic transaminase, a typical B6-enzyme of mammalian tissues. The results obtained were, however, rather contrary to expectation; no inhibition by berberine could be observed in spite of many attempts. It appears, therefore, that the susceptibility of the mammalian 6 enzyme is significantly different from that of bacterial 6 enzymes for reasons to be elucidated in future.

Inhibitory effect of cycloserine on some enzymic activities related to vitamin B6.

It is considered that cycloserine inhibits the vitamin B6-enzyme system in bacteria by forming a complex with vitamin B6.