Vertebrate Opsins Belonging to Different Classes Vary in Constitutively Active Properties Resulting from Salt-Bridge Mutations
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- 作者：Benjamin Nickle ; Susan E. Wilkie ; Jill A. Cowing ; David M. Hunt ; and Phyllis R. Robinson
- 刊名：Biochemistry
- 出版年：2006
- 出版时间：June 13, 2006
- 年：2006
- 卷：45
- 期：23
- 页码：7307 - 7313
- 全文大小：203K
- 年卷期：v.45,no.23(June 13, 2006)
- ISSN：1520-4995
文摘
Vertebrate opsins are classified into one of five classes on the basis of amino acid similarity.These classes are short wavelength sensitive 1 and 2 (SWS1, SWS2), medium/long wavelength sensitive(M/LWS), and rod opsin like 1 and 2 (RH1, RH2). In bovine rod opsin (RH1), two critical amino acidsform a salt bridge in the apoprotein that maintains the opsin in an inactive state. These residues are K296,which functions as the chromophore binding site, and E113, which functions as the counterion to theprotonated Schiff base. Corresponding residues in each of the other vertebrate opsin classes are believedto play similar roles. Previous reports have demonstrated that mutations in these critical residues result inconstitutive activation of transducin by RH1 class opsins in the absence of chromophore. Additionally,recent reports have shown that an E113Q mutation in SWS1 opsin is constitutively active. Here we askif the other classes of vertebrate opsins maintain activation characteristics similar to that of bovine RH1opsin. We approach this question by making the corresponding substitutions which disrupt the K296/E113 salt bridge in opsins belonging to the other vertebrate opsin classes. The mutant opsins are testedfor their ability to constitutively activate bovine transducin. We demonstrate that mutations disruptingthis key salt bridge produce constitutive activation in all classes. However, the mutant opsins differ intheir ability to be quenched in the dark state by the addition of chromophore as well as in their level ofconstitutive activation. The differences in constitutive activation profiles suggest that structural differencesexist among the opsin classes that may translate into a difference in activation properties.