Subcloning, characterization, and affinity labeling of Escherichia coli glycinamide ribonucleotide transformylase
- 13 February 1990
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 29 (6) , 1436-1443
- https://doi.org/10.1021/bi00458a014
Abstract
Glycinamide ribonucleotide transformylase (GAR TFase; EC 2.1.2.2) has been purified 70-fold to apparent homogeneity from Escherichia coli harboring and expression vector encoding the purN gene product, Gar TFase. The protein is a monomer of Mr 23 241 and catalyzes a single reaction. Steady-state kinetic parameters for the enzyme have been obtained. The structural requirements for cofactor utilization have been investigated and found to parallel those of the multifunctional avian enzyme. The enzyme was inactivated with the affinity label N10-(bromoacetyl)-5,8-dideazafolate in a stiochiometric and active site-specific manner. The ionization state of cofactor analogue in the enzyme-cofactor complex appears to require the dissociation of the proton at N3 of the pyrimidine within the complex.This publication has 25 references indexed in Scilit:
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