Slow binding of retinal to rhodopsin mutants G90D and T94D.
In an examination of the effect of three rhodopsin night blindness mutations on the rate of association of 11-cis-retinal with opsin, one of the mutations (G90D) was found to slow the rate of reaction by more than 80-fold. This effect does not appear to be general to night blindness mutations as the two other mutants (A292E and T94I) were not found to bind retinal with slowed kinetics. However, T94D was similar to G90D in that the rate of retinal binding was dramatically slowed. Gly90 and Thr94 are both located in the active site of the protein close to the Schiff base counterion Glu113. Thus, the slow kinetics of Schiff base formation appear to correlate with the introduction of a negative charge close to the Schiff base counterion, suggesting a possible role for Glu113 as a catalytic base in this reaction. Consistent with this model, the E113Q mutant was also found to bind retinal more slowly than the wild type.
Gross
Alecia K
AK
Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02454, USA.
Xie
Guifu
G
Oprian
Daniel D
DD
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000235:genetics*; D018698:Glutamic Acid / Q000235:genetics; D005973:Glutamine / Q000235:genetics; D005998:Glycine / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009755:Night Blindness / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D011489:Protein Denaturation; D012172:Retinaldehyde / Q000737:chemistry*; D012243:Rhodopsin / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D012545:Schiff Bases / Q000737:chemistry; D013056:Spectrophotometry, Ultraviolet; D013912:Threonine / Q000235:genetics*
D012545:Schiff Bases; D005973:Glutamine; D013912:Threonine; D001224:Aspartic Acid; D018698:Glutamic Acid; D012243:Rhodopsin; D000409:Alanine; D012172:Retinaldehyde; D005998:Glycine