scholarly journals Assay and properties of diaminopimelate epimerase from Bacillus megaterium

1969 ◽  
Vol 113 (4) ◽  
pp. 589-601 ◽  
Author(s):  
P. J. White ◽  
B. Lejeune ◽  
Elizabeth Work
Keyword(s):  
Bacillus Megaterium ◽  
Vitamin B6 ◽  
Pyridoxal Phosphate ◽  
Initial Rate ◽  
Equilibrium Mixture ◽  
Enzymic Activity ◽  
Diaminopimelic Acid ◽  
Michaelis Constants ◽  
Assay Methods ◽  
Derivatives Of

1. Diaminopimelate epimerase from a soluble extract of Bacillus megaterium N.C.I.B. 7581 was purified about 25-fold by fractionation with ammonium sulphate and chromatography on calcium phosphate gel–cellulose. The product was impure but was unstable on further purification. 2. Quantitative assay methods for the enzyme were devised in which meso- or ll-diaminopimelic acid may be the substrate. 3. Between 25° and 45° at pH7·0 enzyme action leads to an equilibrium mixture containing 65% meso-isomer and 35% ll-isomer. 4. The initial rate of epimerization was 2–3 times as fast with ll-diaminopimelic acid as substrate as with the meso-isomer; a number of other amino acids were not racemized by the enzyme. The Michaelis constants at 37° were 6·7mm (ll-isomer) and 100mm (meso-isomer); with both substrates enzyme activity was maximal at pH7–8. The relative rates of epimerization of ll-diaminopimelic acid at 25°, 37° and 45° were 0·77:1·00:1·15. 5. A thiol compound (of which 2,3-dimercaptopropan-1-ol was the most effective) was needed as an activator of the purified enzyme. 6. Carbonylbinding reagents and several other compounds did not inhibit diaminopimelate epimerase. 7. Pyridoxal phosphate did not stimulate enzymic activity even in preparations that had been almost completely freed of derivatives of vitamin B6 (as shown by microbiological assay).

scholarly journals The Affinity of Carboplatin to B-Vitamins and Nucleobases

2021 ◽  
Vol 22 (7) ◽  
pp. 3634
Author(s):  
Beata Szefler ◽  
Przemysław Czeleń ◽  
Przemysław Krawczyk
Keyword(s):  
Vitamin B6 ◽  
Pyridoxal Phosphate ◽  
Antitumor Agents ◽  
Lone Pair ◽  
Chemical Compounds ◽  
Free Energies ◽  
Aromatic Rings ◽  
Anti Cancer ◽  
Potential Impact ◽  
Reactive Molecule

Platinum compounds have found wide application in the treatment of various types of cancer and carboplatin is one of the main platinum-based drugs used as antitumor agents. The anticancer activity of carboplatin arises from interacting with DNA and inducing programmed cell death. However, such interactions may occur with other chemical compounds, such as vitamins containing aromatic rings with lone-pair orbitals, which reduces the anti-cancer effect of carboplatin. The most important aspect of the conducted research was related to the evaluation of carboplatin affinity to vitamins from the B group and the potential impact of such interactions on the reduction of therapeutic capabilities of carboplatin in anticancer therapy. Realized computations, including estimation of Gibbs Free Energies, allowed for the identification of the most reactive molecule, namely vitamin B6 (pyridoxal phosphate). In this case, the computational estimations indicating carboplatin reactivity were confirmed by spectrophotometric measurements.

COMMITTEE ON NUTRITION

PEDIATRICS ◽  
1966 ◽  
Vol 38 (6) ◽  
pp. 1068-1076
Author(s):  
Keyword(s):  
Vitamin B6 ◽  
Pyridoxal Phosphate ◽  
Normal Population ◽  
Clinical Signs ◽  
Normal Activity ◽  
Limited Information ◽  
Adequate Intake ◽  
Time Data ◽  
Binding Properties ◽  
Adequate Diet

Because of the limited information available it is not possible to derive precise figures for daily requirements of vitamin B6 in infants and children at this time. Data currently available suggest that the daily need in childhood is 0.5 to 1.5 mg and in adolescence is 1.5 to 2 mg. The requirement in infancy is clearly related to protein intake and is 20µg/gm of dietary protein. Requirements of a few individuals will undoubtedly be higher than the estimates for the normal population. Some of these patients will manifest frank biochemical and clinical signs of deficiency which will usually be promptly reversed by administration of small additional amounts of pyridoxine. Another group of patients will require large amounts of the vitamin to balance the heritable alteration in binding properties of a specific apoenzyme requiring pyridoxal phosphate for normal activity. It would appear that most infants, children and adults will have little difficulty in achieving an adequate intake of vitamin B6 if they receive what is considered to be in other respects an adequate diet.

scholarly journals Ligand binding on to maize (Zea mays) malate synthase: a structural study

1994 ◽  
Vol 303 (2) ◽  
pp. 413-421 ◽  
Author(s):  
S Beeckmans ◽  
A S Khan ◽  
L Kanarek ◽  
E Van Driessche
Keyword(s):  
Zea Mays ◽  
Ligand Binding ◽  
Conformational Change ◽  
Limited Proteolysis ◽  
Enzymic Activity ◽  
Malate Synthase ◽  
Acetyl Coa ◽  
Original Activity ◽  
Kinetic Measurements ◽  
Michaelis Constants

A kinetic and ligand binding study on maize (Zea mays) malate synthase is presented. It is concluded from kinetic measurements that the enzyme proceeds through a ternary-complex mechanism. Michaelis constants (Km,glyoxylate and Km,acetyl-CoA) were determined to be 104 microM and 20 microM respectively. C.d. measurements in the near u.v.-region indicate that a conformational change is induced in the enzyme by its substrate, glyoxylate. From these studies we are able to calculate the affinity for the substrate (Kd,glyoxylate) as 100 microM. A number of inhibitors apparently trigger the same conformational change in the enzyme, i.e. pyruvate, glycollate and fluoroacetate. Another series of inhibitors bearing more bulky groups and/or an extra carboxylic acid also induce a conformational change, which is, however, clearly different from the former one. Limited proteolysis with trypsin results in cleavage of malate synthase into two fragments of respectively 45 and 19 kDa. Even when no more intact malate synthase chains are present, the final enzymic activity still amounts to 30% of the original activity. If trypsinolysis is performed in the presence of acetyl-CoA, the cleavage reaction is appreciably slowed down. The dissociation constant for acetyl-CoA (Kd,acetyl-CoA) was calculated to be 14.8 microM when the glyoxylate subsite is fully occupied by pyruvate and 950 microM (= 50 x Km) when the second subsite is empty. It is concluded that malate synthase follows a compulsory-order mechanism, glyoxylate being the first-binding substrate. Glyoxylate triggers a conformational change in the enzyme and, as a consequence, the correctly shaped binding site for acetyl-CoA is created. Demetallization of malate synthase has no effect on the c.d. spectrum in the near u.v.-region. Moreover, glyoxylate induces the same spectral change in the absence of Mg2+ as in its presence. Nevertheless, malate synthase shows no activity in the absence of the cation. We conclude that Mg2+ is essential for catalysis, rather than for the structure of the enzyme's catalytic site.

Enzymic activity and .gamma.-phosphorus-32 labeling of fluorescent derivatives of cytidine triphosphate and adenosine triphosphate

1973 ◽  
Vol 95 (3) ◽  
pp. 961-962 ◽  
Author(s):  
Jorge R. Barrio ◽  
Laurence G. Dammann ◽  
Leslie H. Kirkegaard ◽  
Robert L. Switzer ◽  
Nelson J. Leonard
Keyword(s):  
Adenosine Triphosphate ◽  
Enzymic Activity ◽  
Derivatives Of ◽  
Fluorescent Derivatives

scholarly journals The action on cellulose and its derivatives of a purified 1,4-β-glucanase from Trichoderma koningii

1981 ◽  
Vol 199 (2) ◽  
pp. 409-417 ◽  
Author(s):  
G Halliwell ◽  
R Vincent
Keyword(s):  
Specific Activity ◽  
High Specificity ◽  
Enzymic Activity ◽  
Transferase Activity ◽  
Beta Glucanase ◽  
Trichoderma Koningii ◽  
Initial Substrate ◽  
Central Bond ◽  
Beta Glucosidase ◽  
Derivatives Of

The specific properties have been examined of the 1,4-beta-glucanase component of Trichoderma koningii that participates in an early and effective stage of random breakdown of native cellulose to short fibres. The enzyme was purified and freed from associated components of the cellulase complex (particularly beta-glucosidase) that interfere with, and complicate interpretation of, the action of such enzymes. Purification increased the specific activity 25-fold over culture filtrates; the enzyme hydrolysed CM-cellulose faster than the purified beta-glucosidase from the same organism hydrolysed any of its substrates (cellobiose or cellodextrins). The specificity of the glucanase was directed towards soluble derivatives of cellulose, CM-cellulose and cellodextrins, and not to insoluble cellulose or alpha-linked polymers. The approximate Km was 2.5 mg of CM-cellulose . ml-1 at 37 degrees C at the optimum pH, 5.5, where enzymic activity was maximal with 6--7 mg of CM-cellulose . ml-1 and inhibited by higher concentrations. The temperature optimum was 60 degrees C. The glucanase attacked larger cellodextrins (cellohexaose to cellotetraose, in that order) much more readily than smaller dextrins (cellobiose and cellotriose) and released a mixture of products, glucose up to cellopentaose, which was quantitatively determined after chromatography on charcoal. Similar examination of hydrolysates of the reduced cellodextrins showed clearly the high specificity of the enzyme for the central bond of its natural substrates (the cellodextrins), whatever their chain length, and indicated the nature of the enzyme as an endoglucanase. Outer bonds shared a weaker, but similar, susceptibility to enzymic cleavage. Transferase activity was absent and no larger dextrins than the initial substrate were formed.

scholarly journals The Alkaline Phosphatase (ALPL) Locus Is Associated with B6 Vitamer Levels in CSF and Plasma

Genes ◽  
2018 ◽  
Vol 10 (1) ◽  
pp. 8 ◽  
Author(s):  
Loes Loohuis ◽  
Monique Albersen ◽  
Simone de Jong ◽  
Timothy Wu ◽  
Jurjen Luykx ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Vitamin B6 ◽  
Pyridoxal Phosphate ◽  
Genome Wide Association Study ◽  
Active Form ◽  
Minor Allele ◽  
Chromosome 1 ◽  
Joint Analysis ◽  
Genome Wide ◽  
A Genome

The active form of vitamin B6, pyridoxal phosphate (PLP), is essential for human metabolism. The brain is dependent on vitamin B6 for its neurotransmitter balance. To obtain insight into the genetic determinants of vitamin B6 homeostasis, we conducted a genome-wide association study (GWAS) of the B6 vitamers pyridoxal (PL), PLP and the degradation product of vitamin B6, pyridoxic acid (PA). We collected a unique sample set of cerebrospinal fluid (CSF) and plasma from the same healthy human subjects of Dutch ancestry (n = 493) and included concentrations and ratios in and between these body fluids in our analysis. Based on a multivariate joint analysis of all B6 vitamers and their ratios, we identified a genome-wide significant association at a locus on chromosome 1 containing the ALPL (alkaline phosphatase) gene (minimal p = 7.89 × 10−10, rs1106357, minor allele frequency (MAF) = 0.46), previously associated with vitamin B6 levels in blood. Subjects homozygous for the minor allele showed a 1.4-times-higher ratio between PLP and PL in plasma, and even a 1.6-times-higher ratio between PLP and PL in CSF than subjects homozygous for the major allele. In addition, we observed a suggestive association with the CSF:plasma ratio of PLP on chromosome 15 (minimal p = 7.93 × 10−7, and MAF = 0.06 for rs28789220). Even though this finding is not reaching genome-wide significance, it highlights the potential of our experimental setup for studying transport and metabolism across the blood–CSF barrier. This GWAS of B6 vitamers identifies alkaline phosphatase as a key regulator in human vitamin B6 metabolism in CSF as well as plasma. Furthermore, our results demonstrate the potential of genetic studies of metabolites in plasma and CSF to elucidate biological aspects underlying metabolite generation, transport and degradation.

The relationship between plasma albumin, alkaline phosphatase and pyridoxal phosphate concentrations in plasma and red cells: Implications for assessing vitamin B6 status

2020 ◽  
Vol 39 (9) ◽  
pp. 2824-2831 ◽  
Author(s):  
Dinesh Talwar ◽  
Anthony Catchpole ◽  
John M. Wadsworth ◽  
Barry J. Toole ◽  
Donald C. McMillan
Keyword(s):  
Alkaline Phosphatase ◽  
Vitamin B6 ◽  
Pyridoxal Phosphate ◽  
Red Cells ◽  
Plasma Albumin ◽  
The Relationship
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