8VSX

NMR Structure of GCAP5 R22A

Summary for 8VSX

• Entry DOI: 10.2210/pdb8vsx/pdb
• NMR Information: BMRB: 51784
• Descriptor: Guanylyl cyclase-activating protein 1 (1 entity in total)
• Functional Keywords: ef-hand, gcap5, phototransduction, retinal guanylyl cyclase, metal binding protein
• Biological source: Danio rerio (zebrafish)
• Total number of polymer chains: 1
• Total formula weight: 22403.27

Authors

Cudia, D.L.,Ames, J.B. (deposition date: 2024-01-24, release date: 2024-05-08, Last modification date: 2024-06-05)

Primary citation

Cudia, D.L.,Ahoulou, E.O.,Bej, A.,Janssen, A.N.,Scholten, A.,Koch, K.W.,Ames, J.B.
NMR Structure of Retinal Guanylate Cyclase Activating Protein 5 (GCAP5) with R22A Mutation That Abolishes Dimerization and Enhances Cyclase Activation.
Biochemistry, 63:1246-1256, 2024

PubMed Abstract: Guanylate cyclase activating protein-5 (GCAP5) in zebrafish photoreceptors promotes the activation of membrane receptor retinal guanylate cyclase (GC-E). Previously, we showed the R22A mutation in GCAP5 (GCAP5) abolishes dimerization of GCAP5 and activates GC-E by more than 3-fold compared to that of wild-type GCAP5 (GCAP5). Here, we present ITC, NMR, and functional analysis of GCAP5 to understand how R22A causes a decreased dimerization affinity and increased cyclase activation. ITC experiments reveal GCAP5 binds a total of 3 Ca, including two sites in the nanomolar range followed by a single micromolar site. The two nanomolar sites in GCAP5 were not detected by ITC, suggesting that R22A may affect the binding of Ca to these sites. The NMR-derived structure of GCAP5 is overall similar to that of GCAP5 (RMSD = 2.3 Å), except for local differences near R22A (Q19, W20, Y21, and K23) and an altered orientation of the C-terminal helix near the N-terminal myristate. GCAP5 lacks an intermolecular salt bridge between R22 and D71 that may explain the weakened dimerization. We present a structural model of GCAP5 bound to GC-E in which the R22 side-chain contacts exposed hydrophobic residues in GC-E. Cyclase assays suggest that GC-E binds to GCAP5 with ∼25% higher affinity compared to GCAP5, consistent with more favorable hydrophobic contact by R22A that may help explain the increased cyclase activation.

PubMed: 38662574
DOI: 10.1021/acs.biochem.4c00046

Experimental method

SOLUTION NMR