Kinetic relationships between the various activities of the formyl-methenyl-methylenetetrahydrofolate synthetase.
The formyl-methenyl-methylenetetrahydrofolate synthetase from chicken liver catalyzes the formation of the 10-formyl- and 5,10-methenyltetrahydrofolate cofactors via three enzymatic activities. In this report we define the kinetic relationships between the activities of this trifunctional protein. An investigation of the time course for 10-formyl cofactor synthesis by computer modeling indicates that commencing with tetrahydropteroyltriglutamate, the activities of the synthetase/cyclohydrolase couple act as separate enzymic species. In contrast, 10-formyl cofactor formation from the 5,10-methylene cofactor utilizing the dehydrogenase/cyclohydrolase couple is described by a single or interactive site model that partitions the 5,10-methenyl intermediate primarily (85%) to the 10-formyl product. An unusual characteristic of the latter coupled activities is the negligible cyclohydrolase activity toward exogenous 5,10-methenyl cofactor, which serves as substrate in the individual activity assay. This is based on (1) competitive inhibition by 5,11-methenyltetrahydrohomofolate against the 5,10-methenyl derivative in the cyclohydrolase-catalyzed hydrolysis but the absence of such inhibition in the dehydrogenase/cyclohydrolase couple and (2) a pulse-chase experiment showing the failure of chase 5,10-methenyl cofactor to dilute the 10-formyl product derived from the coupled activities. The result of this coupling is to minimize the concentration of the 5,10-methenyl species, consistent with its noninvolvement in de novo purine biosynthesis.
D000619:Aminohydrolases / Q000378:metabolism*; D000818:Animals; D002645:Chickens; D005492:Folic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005574:Formate-Tetrahydrofolate Ligase / Q000378:metabolism*; D007700:Kinetics; D002955:Leucovorin / Q000378:metabolism; D008025:Ligases / Q000378:metabolism*; D008099:Liver / Q000201:enzymology; D008433:Mathematics; D045064:Methenyltetrahydrofolate Cyclohydrolase; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP) / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism*; D013763:Tetrahydrofolates / Q000378:metabolism; D013997:Time Factors
D009097:Multienzyme Complexes; D013763:Tetrahydrofolates; C100143:tetrahydrohomofolic acid; C021836:formyl-methenyl-methylenetetrahydrofolate synthetase; C018092:5,10-methenyltetrahydrofolate; C032418:5,11-methenyltetrahydrohomofolate; D005492:Folic Acid; D010088:Oxidoreductases; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP); D000619:Aminohydrolases; D045064:Methenyltetrahydrofolate Cyclohydrolase; D008025:Ligases; D005574:Formate-Tetrahydrofolate Ligase; D002955:Leucovorin