TY - JOUR
T1 - Transmembrane segment IV contributes a functionally important interface for oligomerization of the class II G protein-coupled secretin receptor
AU - Harikumar, Kaleeckal G.
AU - Pinon, Delia I.
AU - Miller, Laurence J.
N1 - Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.
PY - 2007/10/19
Y1 - 2007/10/19
N2 - Oligomerization of the Class II G protein-coupled secretin receptor has been reported, but the molecular basis for this and its functional significance have not been determined. In the current work, we have examined the possible contribution of each of the transmembrane (TM) segments of this receptor to its homo-oligomerization, using the method of competitive disruption screening for inhibition of receptor bioluminescence resonance energy transfer signal. TM IV was the only segment that was found to disrupt receptor bioluminescence resonance energy transfer. Evaluation of predicted interhelical and lipid-exposed faces of this TM segment demonstrated that its lipid-exposed face represented the determinant for oligomerization. This was further confirmed by mutagenesis of the intact secretin receptor. Morphological FRET was utilized to demonstrate that secretin receptor oligomerization occurred at the cell surface and that this oligomerization was disrupted by mutating Gly243 and Ile247, key residues within the lipid-exposed face of TM IV. Although disruption of the receptor oligomerization interface had no effect on secretin binding parameters, it reduced the ability of secretin to stimulate intracellular cAMP. This supports a clear functional effect of oligomerization of this receptor. Such an effect might be particularly relevant to clinical situations in which this receptor is overexpressed, such as in certain neoplasms.
AB - Oligomerization of the Class II G protein-coupled secretin receptor has been reported, but the molecular basis for this and its functional significance have not been determined. In the current work, we have examined the possible contribution of each of the transmembrane (TM) segments of this receptor to its homo-oligomerization, using the method of competitive disruption screening for inhibition of receptor bioluminescence resonance energy transfer signal. TM IV was the only segment that was found to disrupt receptor bioluminescence resonance energy transfer. Evaluation of predicted interhelical and lipid-exposed faces of this TM segment demonstrated that its lipid-exposed face represented the determinant for oligomerization. This was further confirmed by mutagenesis of the intact secretin receptor. Morphological FRET was utilized to demonstrate that secretin receptor oligomerization occurred at the cell surface and that this oligomerization was disrupted by mutating Gly243 and Ile247, key residues within the lipid-exposed face of TM IV. Although disruption of the receptor oligomerization interface had no effect on secretin binding parameters, it reduced the ability of secretin to stimulate intracellular cAMP. This supports a clear functional effect of oligomerization of this receptor. Such an effect might be particularly relevant to clinical situations in which this receptor is overexpressed, such as in certain neoplasms.
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U2 - 10.1074/jbc.M702325200
DO - 10.1074/jbc.M702325200
M3 - Article
C2 - 17726027
AN - SCOPUS:35649028687
SN - 0021-9258
VL - 282
SP - 30363
EP - 30372
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 42
ER -