A new member of alpha 1-adrenoceptor-coupled G alpha h (transglutaminase II) family in pig heart: purification and characterization.
We previously reported an identification of a 77-kDa GTP-binding protein that co-purified with the alpha 1-adrenoceptor following ternary complex formation. In the present paper, we report on the purification and characterization of this GTP-binding protein (termed G alpha h5) isolated from pig heart membranes. After solubilization of pig heart membranes with NaCl, G alpha h5 was purified by sequential chromatographies using DEAE-Cellulose, Q-Sepharose, and GTP-agarose columns. The protein displayed high-affinity GTP gamma S binding which is Mg(2+)-dependent and saturable. The relative order of affinity of nucleotide binding by G alpha h5 was GTP > GDP > ITP >> ATP > or = adenyl-5'-yl imidodiphosphate, which was similar to that observed for other heterotrimeric G-proteins involved in receptor signaling. Moreover, the G alpha h5 demonstrated transglutaminase (TGase) activity that was blocked either by EGTA or GTP gamma S. In support of these observations, the G alpha h5 was recognized by a specific antibody to G alpha h7 or TGase II, indicating a homology with G alpha h (TGase II) family. These results demonstrate that 77-kDa G alpha h5 from pig heart is an alpha 1-adrenoceptor-coupled G alpha h (TGase II) family which has species-specificity in molecular mass.
Yoo
S M
SM
Department of Biochemistry, College of Medicine, Chung-Ang University, Seoul, Korea.
Jeong
H S
HS
Han
K J
KJ
Cho
S H
SH
Lee
H S
HS
Yun
H Y
HY
Kwon
N S
NS
Baek
K J
KJ
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D003429:Cross Reactions; D019204:GTP-Binding Proteins / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D008970:Molecular Weight; D009206:Myocardium / Q000737:chemistry*; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism; D013552:Swine; D011503:Transglutaminases / Q000378:metabolism
D018340:Receptors, Adrenergic, alpha-1; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D011503:Transglutaminases; D019204:GTP-Binding Proteins