Purification and characterization of dihydrofolate reductase from Lactobacillus leichmannii.
Indian J Biochem Biophys
;
2000 Apr; 37(2): 121-9
Article
in English
| IMSEAR
| ID: sea-27218
ABSTRACT
Dihydrofolate reductase (DHFR) (5,6,7,8-THF NADDP+ oxidoreductase, EC 1.5.1.3) was purified 205-fold to apparent homogeneity from the crude extracts of Lactobacillus leichmannii. It has UV absorption maxima at 280 nm, M(r) of 20,000, Stokes radius of 0.34 nm and a S20.w value of 0.12 S. The preparation showed the presence of 168 amino acid residues with threonine and lysine as the NH2- and COOH- terminal end-groups respectively and a single reactive sulfhydryl group. pCMB inhibited the enzyme activity (IC50 = 2 microM). The enzyme has a pH optimum of 7.4 and is thermally inactivated at > 35 degrees C. It is activated by 0.1 M KCl and KI and 2 M urea. 3-4 M urea completely inactivated the enzyme. Enzyme has Km values of 3.5 microM and 6.2 microM for NADPH and DHF respectively, and a Ki value of 7 nM for MTX, the inhibition being competitive.
Full text:
Available
Index:
IMSEAR (South-East Asia)
Main subject:
Tetrahydrofolate Dehydrogenase
/
Kinetics
/
Amino Acids
/
Lactobacillus
/
Molecular Weight
Language:
English
Journal:
Indian J Biochem Biophys
Year:
2000
Type:
Article
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