Purification and characterization of a feline hepatic insulin receptor.
OBJECTIVE
To elucidate the functional characteristics of a highly purified soluble liver insulin receptor in cats.


METHODS
Frozen livers from domestic cats were obtained commercially.


METHODS
The feline hepatic insulin receptor was purified from Triton X-100 solubilized plasma membranes by the use of several chromatography matrices, including affinity chromatography on an insulin-Sepharose matrix.


RESULTS
The receptor, although not homogeneous, was purified 3,000-fold. Two silver-stained protein bands were identified following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with molecular weight of 134,000 and 97,000, which are similar to insulin receptors isolated from other animals. This isolated receptor had steady-state insulin binding by 40 minutes at 24 C. Optimal insulin binding occurred at pH 7.8 and with 150 mM NaCl. Under these conditions, a curvilinear Scatchard plot was obtained with the isolated receptor. Using a 2 binding-site model, the feline insulin receptor had a high-affinity low-capacity site with a dissociation constant (KD; nM) of 3 and a low-affinity high-capacity site with a K(D) of 1,180. The receptor also had tyrosine kinase activity toward an exogenous substrate that was stimulated by insulin and protamine.


CONCLUSIONS
Many of the reported characteristics of the liver insulin receptor in cats are similar to those for the receptor isolated from other animals and tissues, although some differences exist. These similarities suggest that characterization of the feline insulin receptor is important to understanding insulin resistance in cats with diabetes as well as in humans with diabetes.
Paxton
R
R
Department of Anatomy, Physiology & Pharmacology, College of Veterinary Medicine, Auburn University, AL 36849, USA.
Ye
L X
LX
D000818:Animals; D000829:Animals, Domestic; D001665:Binding Sites; D002415:Cats; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007328:Insulin / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000737:chemistry* / Q000378:metabolism*; D008970:Molecular Weight; D011972:Receptor, Insulin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
D007328:Insulin; D011972:Receptor, Insulin