title: Binary and ternary complexes of dihydrofolate reductase with substrates, coenzymes and inhibitors. Circular dichroic and magnetic circular dichroic studies.
abstract: Interaction of several representative folate, quinazoline and pyridine nucleotide derivatives with dihydrofolate reductase from amethopterin-resistant Lactobacillus casei induces dramatic changes in its circular dichroic spectral properties. The binding of dihydrofolate induces a large extrinsic Cotton effect at 295 nm ([theta] = 113 800 deg . cm2 . dm-1). The generation of this band by dihydrofolate is strictly dependent on complex formation with a single substrate binding site and a KD = 7 . 10(-6) M. The other binary complexes examined include the enzyme . NADPH, enzyme . amethopterin, enzyme . folate, and enzyme . methasquin. All such complexes differ in spectral detail, the negative ellipticity at 330 nm being characteristic of the "folate site" complexes. The circular dichroic spectrum of the ternary complex of reductase . NADPH . methotrexate shows a positive symmetrical band centered at 360 nm ([theta] - 32 000 deg . cm2 . dm-1). Since both of the corresponding binary complexes exhibit negative bands in this region, this induced band represents a unique molecular property of the ternary complex. Chemical modification of a single tryptophan residue of the enzyme, as determined from magnetic circular dichroism spectra, results in a complete loss in the ability to bind either dihydrofolate or NADPH.
mesh_terms: D001665:Binding Sites; D002942:Circular Dichroism; D007700:Kinetics; D007780:Lacticaseibacillus casei / Q000201:enzymology; D008280:Magnetics; D011485:Protein Binding; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism
chemical_list: D013762:Tetrahydrofolate Dehydrogenase