共查询到20条相似文献,搜索用时 9 毫秒
1.
Kenneth R. Hanson 《Archives of biochemistry and biophysics》1981,211(2):575-588
The binding of substrate and product analogs to phenylalanine ammonia-lyase (EC 4.3.1.5) from maize has been studied by a protection method. The ligand dissociation constants, KL, were estimated from the variation with [L] of the pseudo-first-order rate constants for enzyme inactivation by nitromethane. The phenylalanine analogs d- and l-2-aminooxy-3-phenylpropionic acid showed KL, values over 20,000-fold lower than the Km for l-phenylalanine. From these and other KL values it is deduced that when the enzyme binds l-phenylalanine the structural free energy stored in the protein is higher than when it binds the superinhibitors. Models for binding d- and l-phenylalanine and the superinhibitors are described. The enantiomeric pairs are considered to have similar KL values because they pack into the active site in a mirror-image relationship. If the elimination reaction approximates to the least-motion course deduced on stereoelectronic grounds, the mirror-image packing of the superinhibitors into the active site mimics the conformation inferred for a transition state in the elimination. It appears, therefore, that structural changes take place in the enzyme as the transition state conformation is approached causing stored free energy to be released. This lowers the activation free energy for the elimination reaction and accounts for the strong binding by the above analogs. 相似文献
2.
The uptake of l-DOPA (l-3,4-dihydroxyphenylalanine) was studied in normal human red blood cells in vitro using l-[3-14C]DOPA. Uptake was slow, tending towards a distribution ratio close to unity with a half-time to equilibrium of one hour. Uptake was not Na+-dependent. Concentration dependence studies showed both saturable and non-saturable components of uptake, and inhibition studies using l-leucine and l-tryptophan suggest that the L and T systems of red cell amino acid uptake are involved. A powerful inhibitor of both systems, 3,4-dihydroxy-2-methylpropriophenone (U-0521), is described. It is concluded that uptake is by carrier-mediated facilitated diffusion via the L and T systems for which l-DOPA has low affinity. 相似文献
3.
Heinz Rennenberg 《Phytochemistry》1983,22(7):1557-1560
Cysteine desulfhydrase activity in leaf discs of cucurbit plants is enhanced 2–4-fold by preincubation with l or d-cysteine. Preincubation with structural analogs of cysteine also stimulated the activity of the enzyme, but to a smaller extent. Maximal increase in cysteine desulfhydrase activity was observed by preincubation with 5 mM or higher concentrations of cysteine. Although not caused by activation, stimulation of the enzyme activity was half-maximal within less than 15 min. Whereas the increase in cysteine desulfhydrase activity by preincubation of leaf discs with cysteine was light independent, pretreatment of the entire plant with light or dark determined the leaf discs' potential for stimulation of the enzyme. Exposure to darkness for 4 hr reduced this potential by 60%. It is concluded that the potential for stimulation of cysteine desulfhydrase activity by preincubation with cysteine is regulated by a compound not synthesized, but metabolized, in the leaf tissue. This regulatory compound may be supplied to the leaves by long-distance transport. 相似文献
4.
Carol L. Stahl Gary A. Sojka 《Biochimica et Biophysica Acta (BBA)/General Subjects》1973,297(2):241-245
d-malate replaced l-malate in supporting both photosynthetic (anaerobic, light) and heterotrophic (aerobic, dark) growth of Rhodopseudomonas capsulata. Growth rates and cell yields were nearly equivalent with both enantiomorphs. Addition of glucose to malate culture media increased the growth rate and doubled the cell yield of heterotrophic cultures, but had little effect on photosynthetic cultures. Aerobically-grown cells showed a higher level of substrate-dependent oxygen uptake with l-malate than with d-malate. This preference for l-malate occured even in cells grown on d-malate. No malic racemase activity was detected in extracts of heterotrophically- or photosynthetically-grown cells. 相似文献
5.
Hiroyuki Yamamoto Tadao Hayakawa 《International journal of biological macromolecules》1982,4(5):258-262
The conformations of random and sequential copolypeptides containing l-β-3,4-dihydroxyphenyl-α-alanine (DOPA) and l-glutamic acid (Glu) as well as their protected precursors have been investigated mainly by means of circular dichroism (c.d.) spectroscopy in chloroform or trimethylphosphate as solvent. Protected and deprotected DOPA-containing polypeptides showed a positive ellipticity hand at 285 nm due to the stacked side-chains. The c.d. behaviour depended on the amino acid sequence as well as the amino acid composition. In random copolypeptides, ellipticities versus DOPA content showed a smooth variation, without any sharp changes. This supported the conclusion that poly(DOPA) is a right-handed helix. Although the ellipticities were low compared with the values of a typical α-helix, deprotected DOPA-containing sequential polypeptides are helical from the results of the induced dichroic band at 285 nm and the infrared amide I and II absorption bands. The results obtained were compared with those of sequential polypeptides containing l-tyrosine and l-Glu. 相似文献
6.
Ian D. Hay Thomas M. Annesley Nai S. Jiang Colum A. Gorman 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1981,226(2):383-390
A method for the determination of d- and l-thyroxine in human serum is described. The method involves extraction of thyroxine from serum and the separation of thyroxine enantiomers on a reversed-phase, high-performance liquid chromatographic column by use of a chiral eluent containing l-proline and cupric sulfate. Satisfactory resolution of the enantiomers of thyroxine, triiodothyronine, and reverse triiodothyronine can be achieved in 12 min and, employing amperometric detection to monitor the separation, the detection limit for serum thyroxine is in the range of 1–3 ng per injected sample. 相似文献
7.
In this paper we studied the degradation of l-2-hydroxyglutarate in tissues from rat and man in order to try and find the underlying basis for the accumulation of this metabolite in l-2-hydroxyglutaric patients. The results show that l-2-hydroxyglutarate is not degraded by an oxidase but via a dehydrogenase which was found to be present in liver only. This newly identified enzyme activity was characterized kinetically, although the nature of the reaction product remains to be identified. 相似文献
8.
9.
The reaction of trans(N)-[Co(d-pen)2]− (pen = penicillaminate) with HgCl2 or HgBr2 in the molar ratios of 1:1 gave the sulfur-bridged heterodinuclear complex, [HgX(OH2){Co(d-pen)2}] (X = Cl (1a) or Br (1b)). A similar reaction in the ratio of 2:1 produced the trinuclear complex, [Hg{Co(d-pen)2}2] (1c). The enantiomers of 1a and 1c, [HgCl(OH2){Co(l-pen)2}] (1a′) and [Hg{Co(l-pen)2}2] (1c′), were also obtained by using trans(N)-[Co(l-pen)2]− instead of trans(N)-[Co(d-pen)2]−. Further, the reaction of cis · cis · cis-[Co(d-pen)(l-pen)]− with HgCl2 in the molar ratio of 1:1 resulted in the formation of [HgCl(OH2){Co(d-pen)(l-pen)}] (2a). During the formations of the above six complexes, 1a, 1b, 1c, 1a′, 1c′, and 2a, the octahedral Co(III) units retain their configurations. On the other hand, the reaction of cis · cis · cis-[Co(d-pen)(l-pen)]− with HgCl2 in the molar ratio of 2:1 gave not [Hg{Co(d-pen)(l-pen}2] but [Hg{Co(d-pen)2}{Co(l-pen)2}] (2c), accompanied by the ligand-exchange on the terminal Co(III) units. The X-ray crystal structural analyses show that the central Hg(II) atom in 1c takes a considerably distorted tetrahedral geometry, whereas that in 2c is of an ideal tetrahedron. The interconversion between the complexes is also examined. The electronic absorption, CD, and NMR spectral behavior of the complexes is discussed in relation to the crystal structures of 1c and 2c. 相似文献
10.
The enzymatic decarboxylations of l-DOPA and l-5-hydroxytryptophan (l-5-HTP) by aromatic l-amino acid decarboxylase (AADC) were measured with homogenates from human brain regions, caduate nucleus and hypothalamus, using our new and highly sensitive methods for l-DOPA decarboxylase and l-5-HTP decarboxylase by high-performance liquid chromatography with electrochemical detection (HPLC-ED). Dopamine formed from l-DOPA as substrate was measured for DOPA decarboxylase activity using d-DOPA for the blank. For 5-HTP decarboxylase activity, serotonin (5-HT) formed from l-5-HTP was measured, and the blank value in presence of NSD-1055 was subtracted. NSD-1055 inhibited 5-HTP decarboxylase activity completely at a concentration of 0.2 mM. In this study, the properties of l-5-HTP decarboxylase activity in human caudate nucleus were first examined. AADC activities in human brains were found to be widely variable for both l-DOPA and l-5-HTP as substrates. The ratio of the activities for l-DOPA and l-5-HTP were found to be significantly higher in hypothalamus than in caudate nucleus. AADC activity for l-DOPA in the brain was found to be linear up to 40 min of incubation, while that for l-5-HTP was found to be linear up to 240 min of incubation. The optimum pyridoxal phosphate concentration was found to be similar for both substrates and was between 0.01 and 0.1 mM. The optimum pH values were found to be 7.2 and 8.2 for l-DOPA decarboxylase and l-5-HTP decarboxylase, respectively. Km and Vmax values for a human caudate nucleus l-DOPA decarboxylase were found to be 414 μM and 482 pmol/min/g wet weight, respectively, while those for l-5-HTP decarboxylase were found to be 90 μM and 71 pmol/min/g wet weight, respectively. 相似文献
11.
Murray Goodman Alvin Steinfeld Alan Tonelli Ugo Lepore Manlio Palumbo Bernard Donzel 《Bioorganic chemistry》1974,3(2):184-203
The synthesis of the cyclo-hexadepsipeptide [l-valyl-d-hexahydromandelyl]3 is described. Examination of this macrocyclic compound by 220-MHz nuclear magnetic resonance spectroscopy shows that symmetrical conformations are stabilized in strongly polar solvents (trifluoroacetic acid, acetonitrile), whereas asymmetric conformations are preferred in nonpolar or slightly polar media such as carbon tetrachloride, chloroform, cyclohexane, and benzene.From analysis of the temperature dependence of the chemical shift and of the coupling constants, together with conformational energy calculations, a model is proposed for the preferred conformation of this molecule in nonpolar solvents. 相似文献
12.
Threonine and C1 units from methionine contribute to form an intermediate in the biosynthesis of l-β-(methylenecyclopropyl)-alanine, the toxi 相似文献
13.
Dana W. Aswad 《Analytical biochemistry》1984,137(2):405-409
A sensitive and convenient method for the simultaneous determination of d- and l-aspartic acid in amino acid mixtures is described. The method involves derivatization of the mixture with a chiral fluorogen, followed by high-performance liquid chromatography on a reverse-phase column. The fluorogen used is an adduct of o-phthaldialdehyde with an optically active thiol, N-acetyl-l-cysteine. The sensitivity and accuracy of this method is similar to that using adducts of o-pthaldialdehyde with the achiral thiol, 2-mercaptoethanol. Five picomoles of d-aspartate can be accurately detected in the presence of a 100-fold excess of l-aspartate with a total analysis time (including derivatization) of 10 min. 相似文献
14.
Aminotransferases are pyridoxal phosphate-dependent enzymes whose potential for the biocatalytic production of enantiopure amino acids is increasingly recognized. Because of this, there is a growing interest in engineering them to alter their substrate specificity and to increase their catalytic activity. Here, we report the development of a high-throughput assay for screening α-ketoglutarate-dependent aminotransferase mutant libraries. To achieve this, we exploited the l-glutamate dehydrogenase coupled assay that has previously been shown to allow for aminotransferase activity to be monitored in vitro. We adapted this assay to allow screening of mutant libraries of either l- or d-amino acid specific aminotransferases in a continuous fashion. This assay requiring clarified cell lysates is reproducible, rapid, and sensitive because it allowed for the identification of a catalytically active mutant of Bacillus sp. YM-1 d-amino acid aminotransferase displaying a decrease in kcat/KM of more than two orders of magnitude. In addition, this assay allowed us to discover a mutant of Escherichia coli branched-chain amino acid aminotransferase, F36W, which is approximately 60-fold more specific toward the natural substrate l-leucine than l-phenylalanine as compared with wild type. This result demonstrates the potential of our assay for the discovery of mutant aminotransferases displaying altered substrate specificity, an important goal of enzyme engineering. 相似文献
15.
A simple and rapid technique for the determination of the d-amino acids which are oxidized by d-amino acid oxidase has been presented. This method involves an oxidation of d-amino acids with d-amino acid oxidase in the presence of catalase, and the spectrophotometric determination of the resultant α-keto acids with MBTH. The additions of l-amino acids have no influence on the quantitative estimation of d-amino acids. The method is suitable for the assay of d-amino acids in the presence of the l isomers, and is also applicable for the determination of d-amino acid oxidase activity. 相似文献
16.
The reactions of some 4,6-disulphonates of methyl 2,3-di-O-acyl-(and di-O-methyl)-d-glucopyranosides and -galactopyranosides, with thiocyanate, thioacetate, and thiobenzoate anions, have been studied under a variety of conditions. In the glucoside series, somewhat similar reactivity is shown by isomers differing only in anomeric configuration, and there is no very great difference between the reactivities of a 2,3-dibenzoate and its 2,3-di-O-methyl analogue. In contrast to the situation in the β-d-galactoside series, the presence of O-benzoyl groups in an α-d-galactoside does not have an unfavourable effect on displacement at C-4. Two hexose derivatives containing the novel 4,6-epithio bridge are described. 相似文献
17.
Acid-catalysed condensation of methyl β-d-glycero-l-manno-heptopyranoside with cyclohexanone yielded an approximately 3:1 mixture of the 2,3:6,7- and 2,3:4,7-di-O-cyclohexylideneheptosides (1 and 2), which could be separated either as their benzoates (3 and 4) or as their methyl ethers (5 and 6). The latter compounds afforded the 4- and 6-methyl ethers (7 and 8) of d-glycero-l-manno-heptitol. 相似文献
18.
l-(+)-Glutamate oxidation that is non-pyridine nucleotide dependent is readily carried out by a membrane-bound enzyme in Azotobacter vinelandii strain O. Enzyme activity concentrates in a membranous fraction that is associated with the Azotobacter electron transport system. This l-glutamate oxidation is not dependent on externally added NAD+, NADP+, FAD, or FMN for activity. O2, phenazine methosulfate and ferricyanide all served as relatively good electron acceptors for this reaction; while cytochrome c and nitrotetrazolium blue function poorly in this capacity. Paper chromatographic analyses revealed that the 2,4-dinitrophenylhydrazine derivative formed from the enzymatic oxidation of l-glutamate was α-ketoglutarate, while microdiffusion studies indicated that ammonia was also a key end product. These findings suggest that the overall reaction is an oxidative deamination. Ammonia formation was found to be stoichiometric with the amount of oxygen consumed (2 : 1 respectively, on a molar basis). The oxidation of glutamate was limited to the l-(+)-enantiomer indicating that this reaction is not the generalized type carried out by the l-amino acid oxidase. This oxidoreductase is functionally related to the Azotobacter electron transport system: (a) the activity concentrates almost exclusively in the electron transport fraction; (b) the l-glutamate oxidase activity is markedly sensitive to electron transport inhibitors, i.e. 2-n-heptyl-4-hydroxyquinoline-N-oxide, cyanide, and 4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione; and (c) spectral studies on the Azotobacter R3 fraction revealed that a substantial amount of the flavoprotein (non-heme iron) and cytochrome (a2, a1, b1, c4 and c5) are reduced by the addition of l-glutamate. 相似文献
19.
Lisa Dalla Via Sebastiano Marciani Magno Daniele Marton 《Inorganica chimica acta》2006,359(13):4197-4206
Described herein is the synthesis of two platinum(II) complexes containing l-ornithine (1) or d-ornithine (2) as a ligand. The complexes were obtained by the direct reaction of aqueous solutions of potassium tetrachloroplatinate and l- or d-ornithine. The single crystal structures of 1 and 2 were determined and indicated that the Pt core is surrounded by an almost regular square planar coordination environment. The two complexes were thoroughly characterized by means of FT-IR, FT-Raman and NMR techniques. Furthermore, their ability to inhibit proliferation of human tumor cell lines was tested and the results indicate that 1 and 2 exert different cytotoxic effects. Particularly, the complex with the d-isomer of ornithine (2) showed a significantly greater cytotoxicity than that with the l-isomer (1). Finally, circular dichroism analysis indicated that 1 and 2 interact with DNA in a manner similar to that of cisplatin. 相似文献
20.
The GSH dependence of the metabolic pathways involved in the conversion of cysteine to sulfate in intact cells has been investigated. It was found that hepatocyte-catalysed sulfate formation from added
-cysteine did not occur if hepatocyte GSH was depleted beforehand, but was restored when GSH levels recovered. Furthermore, sulfate formation did not recover in GSH-depleted hepatocytes if GSH synthesis was prevented with buthionine sulfoximine. Thiosulfate formation was, however, markedly enhanced in GSH-depleted hepatocytes. These results suggest that thiosulfate is an intermediate in the formation of inorganic sulfate from
-cysteine and that GSH was required for the conversion of thiosulfate to inorganic sulfate. Much less sulfate was formed if the cysteine was replaced with cysteinesulfinate. Furthermore, sulfate formation from
-cysteine was markedly inhibited by the addition of the transaminase inhibitor
-cycloserine or the γ-cystathionase inhibitor
-propargylglycine. The major routes of sulfate formation from
-cysteine therefore seems to involve pathways that do not involve
-cysteinesulfinate. Similar amounts of sulfate were formed from
-cysteine as
-cysteine. Thiosulfate instead of sulfate was also formed in GSH-depleted hepatocytes. However, sulfate formation from
-cysteine differed from
-cysteine in that it was inhibited by the
-aminoacid oxidase inhibitor sodium benzoate and was not affected by transaminase or γ-cystathionase inhibitors. These results suggest that thiosulfate is an intermediate in sulfate formation from
-cysteine and involves the oxidation of
-cysteine by
-amino acid oxidase to form β-mercaptopyruvate. 相似文献
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