Responses to insulin by two forms of rat hepatic microsomal cytochrome P-450 that undergo major (RLM6) and minor (RLM5b) elevations in diabetes.
Total cytochrome P-450 levels rise in diabetic rats. Two specific forms of cytochrome P-450 that are elevated have been isolated from liver microsomes of streptozotocin-induced idabetic male rats. One enzyme, termed RLM6, metabolizes aniline and acetol, but not testosterone, in a reconstituted system with NADPH-cytochrome P-450 reductase. RLM6 is isolated as a high spin cytochrome with a minimum molecular weight of 53,500. It has a unique amino-terminal amino acid sequence lacking methionine at the amino-terminal position. Polyclonal antibodies to RLM6 recognized most other forms of cytochrome P-450 in Western blots, but could be made monospecific by adsorption to cross-reacting proteins coupled to Sepharose 4B. Using the monospecific antibodies, RLM6 was estimated to be present in microsomes of untreated male rats at 0.04 nmol/mg protein (5% of total P-450). In chronically diabetic rats this level rose to 0.35 nmol/mg protein and 24% of the P-450 content. Immunoreactive protein of molecular weight identical to RLM6 was elevated in microsomes of non-diabetic rats treated with ethanol, acetone, or isoniazid as well as in rats starved for 48 h. Insulin treatment of diabetic rats for 1 week lowered the immunologically detectable levels of RLM6 to levels found in the untreated rat. The other form of cytochrome P-450, RLM5b, does not metabolize aniline and only poorly metabolizes acetol and testosterone. This 52.5-kDa protein is isolated as a predominantly (60%) high spin enzyme. It has a unique NH2-terminal amino acid sequence with methionine as the terminal residue, and is present in untreated male rat liver microsomes at 0.16 nmol/mg protein. It is elevated in diabetes, like RLM6, but treatment with insulin for 1 week does not completely restore the microsomal content to that of the non-diabetic rat.
Favreau
L V
LV
Department of Pharmacology, University of Connecticut Health Center, Farmington 06032.
Malchoff
D M
DM
Mole
J E
JE
Schenkman
J B
JB
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D003577:Cytochrome P-450 Enzyme System / Q000302:isolation & purification / Q000378:metabolism*; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism*; D007328:Insulin / Q000494:pharmacology*; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
D000906:Antibodies; D007328:Insulin; D007527:Isoenzymes; D003577:Cytochrome P-450 Enzyme System