Structural analysis of the complement control protein (CCP) modules of GABA(B) receptor 1a

TY - JOUR

T1 - Structural analysis of the complement control protein (CCP) modules of GABA(B) receptor 1a

T2 - only one of the two CCP modules is compactly folded

AU - Blein, Stanislas

AU - Ginham, Rachel

AU - Uhrin, Dusan

AU - Smith, Brian O

AU - Soares, Dinesh C

AU - Veltel, Stefan

AU - McIlhinney, R A Jeffrey

AU - White, Julia H

AU - Barlow, Paul N

AU - Veltel, Stefan

PY - 2004/11/12

Y1 - 2004/11/12

N2 - The gamma-aminobutyric acid type B (GABA(B)) receptor is a heterodimeric G-protein-coupled receptor. In humans, three splice variants of the GABA(B) receptor 1 (R1) subunit differ in having one, both, or neither of two putative complement control protein (CCP) modules at the extracellular N terminus, prior to the GABA-binding domain. The in vivo function of these predicted modules remains to be discovered, but a likely association with extracellular matrix proteins is intriguing. The portion of the GABA(B) R1a variant encompassing both of its CCP module-like sequences has been expressed, as have the sequences corresponding to each individual module. Each putative CCP module exhibits the expected pattern of disulfide formation. However, the second module (CCP2) is more compactly folded than the first, and the three-dimensional structure of this more C-terminal module (expressed alone) was solved on the basis of NMR-derived nuclear Overhauser effects. This revealed a strong similarity to previously determined CCP module structures in the regulators of complement activation. The N-terminal module (CCP1) displayed conformational heterogeneity under a wide range of conditions whether expressed alone or together with CCP2. Several lines of evidence indicated the presence of native disorder in CCP1, despite the fact that recombinant CCP1 contributes to binding to the extracellular matrix protein fibulin-2. Thus, we have shown that the two CCP modules of GABA(B) R1a have strikingly different structural properties, reflecting their different functions.

AB - The gamma-aminobutyric acid type B (GABA(B)) receptor is a heterodimeric G-protein-coupled receptor. In humans, three splice variants of the GABA(B) receptor 1 (R1) subunit differ in having one, both, or neither of two putative complement control protein (CCP) modules at the extracellular N terminus, prior to the GABA-binding domain. The in vivo function of these predicted modules remains to be discovered, but a likely association with extracellular matrix proteins is intriguing. The portion of the GABA(B) R1a variant encompassing both of its CCP module-like sequences has been expressed, as have the sequences corresponding to each individual module. Each putative CCP module exhibits the expected pattern of disulfide formation. However, the second module (CCP2) is more compactly folded than the first, and the three-dimensional structure of this more C-terminal module (expressed alone) was solved on the basis of NMR-derived nuclear Overhauser effects. This revealed a strong similarity to previously determined CCP module structures in the regulators of complement activation. The N-terminal module (CCP1) displayed conformational heterogeneity under a wide range of conditions whether expressed alone or together with CCP2. Several lines of evidence indicated the presence of native disorder in CCP1, despite the fact that recombinant CCP1 contributes to binding to the extracellular matrix protein fibulin-2. Thus, we have shown that the two CCP modules of GABA(B) R1a have strikingly different structural properties, reflecting their different functions.

KW - Animals

KW - Calcium-Binding Proteins

KW - Calorimetry, Differential Scanning

KW - Circular Dichroism

KW - Disulfides

KW - Extracellular Matrix Proteins

KW - Humans

KW - Models, Molecular

KW - Nuclear Magnetic Resonance, Biomolecular

KW - Peptide Fragments

KW - Protein Binding

KW - Protein Denaturation

KW - Protein Isoforms

KW - Protein Structure, Secondary

KW - Protein Structure, Tertiary

KW - Rats

KW - Receptors, GABA-B

KW - Two-Hybrid System Techniques

U2 - 10.1074/jbc.M406540200

DO - 10.1074/jbc.M406540200

M3 - SCORING: Journal article

C2 - 15304491

VL - 279

SP - 48292

EP - 48306

JO - J BIOL CHEM

JF - J BIOL CHEM

SN - 0021-9258

IS - 46

ER -