Inhibition of dihydrofolate reductase from bacterial and vertebrate sources by folate, aminopterin, methotrexate and their 5-deaza analogues.
The inhibition of dihydrofolate reductases from Escherichia coli and chicken liver by folate, methotrexate, aminopterin and their 5-deaza analogues was investigated to examine the importance of the N-5 nitrogen in slow-binding inhibition. Methotrexate, aminopterin and their 5-deaza analogues acted as slow, tight-binding inhibitors of both enzymes. Inhibition by methotrexate and 5-deazamethotrexate conformed to a mechanism in which there is an initial rapid formation of an enzyme-NADPH-inhibitor complex followed by a slow isomerization of this complex (Mechanism B). Aminopterin exhibited the same type of inhibition with the enzyme from E. coli. With the chicken-liver enzyme, however, the inhibition by aminopterin conformed to another type of slow-binding mechanism which involves only the slow interaction of the inhibitor with the enzyme to form an enzyme-NADPH-inhibitor complex (Mechanism A). The inhibition of both enzymes by 5-deazaaminopterin was also described by Mechanism A. Folate behaved as a classical, steady-state inhibitor of both enzymes, whereas 5-deazafolate exhibited slow-binding inhibition (Mechanism B) with the enzyme from E. coli and classical, steady-state inhibition with the enzyme from chicken liver. The substitution of a carbon for a nitrogen at the 5-position of methotrexate and aminopterin did not affect the tightness of binding of these compounds. By contrast, 5-deazafolate was bound about 4000 times more tightly than folate to the enzyme from E. coli and about 30 times more tightly than folate to the chicken-liver enzyme. Reasons for the differences in the binding of folate and 5-deazafolate are discussed.
D000630:Aminopterin / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D002645:Chickens; D004926:Escherichia coli / Q000201:enzymology*; D005492:Folic Acid / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D005493:Folic Acid Antagonists; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008727:Methotrexate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D011485:Protein Binding; D013329:Structure-Activity Relationship
C040504:5-deazamethotrexate; D005493:Folic Acid Antagonists; C040506:5-deazaaminopterin; C040505:5-deazafolic acid; D005492:Folic Acid; D000630:Aminopterin; D008727:Methotrexate