Spectral changes associated with binding of folate compounds to bacteriophage T4 dihydrofolate reductase.

The difference spectra associated with complex formation at pH 7.0 between bacteriophage T4-specific dihydrofolate reductase and the 4-amino folate analogues, aminopterin, methotrexate, and N1o-formyl-aminopterin, have features which are similar to the difference spectra obtained for the free compounds in acid versus neutral solutions. These results suggest that oxidized 4-amino-folate compounds bind to T4 Hz-folate reductase in the cationic form. This similarity does not extend to the difference spectra for folic acid, Hz-folate, or Hz-aminopterin. The difference spectra of complex formation between Hz-folate and T4 Hz-folate reductase show enhanced absorption with peaks at 305 nm and 243 mu, which are characteristics of the absorption spectra of the pteridine moiety of Ha-folate. We observed no evidence for protonation of Ns of Hz-folate as a result of complex formation. The difference spectrum of the complex between T4 Hz-folate reductase and H+minopterin, but not T4 Hz-folate reductase and aminopterin, is perturbed by the cofactor NADPH. This suggests that the binding site for Hz-aminopterin might not be identical with that for aminopterin.