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Title: Effects of kinase inhibitors and potassium phosphate (KPi) on site-specific phosphorylation of branched chain. cap alpha. -ketoacid dehydrogenase (BCKDH)

BCKDH is phosphorylated by a copurifying kinase at two serine residues on the El..cap alpha.. subunit. Phosphorylation of both sites occurs at about the same rate initially, but inactivation is believed associated only with site 1 phosphorylation. The effects of KPi and known inhibitors of BCKDH kinase, ..cap alpha..-chloroisocaproate (CIC) and branched chain ..cap alpha..-ketoacids (BCKA), on the phosphorylation of purified rat liver BCKDH were studied. Site-specific phosphorylation was quantitated by thin-layer electrophoresis of tryptic peptides followed by densitometric scanning of autoradiograms. Addition of 5 mM KPi was found necessary to stabilize the BCKDH activity at 37/sup 0/C. Increasing the KPi to 50 mM dramatically increased the CIC and BCKA inhibition of site 1 and site 2 phosphorylation. The finding of enhanced sensitivity of inhibitors with 50 mM KPi may facilitate identification of physiologically important kinase effectors. Regardless of the KPi concentration, CIC and the BCKA showed much more effective inhibition of site 2 than site 1 phosphorylation. Although site 1 is the primary inactivating site, predominant inhibition of site 2 phosphorylation may provide a means of modulating kinase/phosphatase control of BCKDH activity under steady state conditions.
Authors:
; ; ;
Publication Date:
OSTI Identifier:
6179702
Report Number(s):
CONF-870644-
Journal ID: CODEN: FEPRA; TRN: 87-034025
Resource Type:
Conference
Resource Relation:
Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States); Journal Volume: 46:6; Conference: 78. annual meeting of the American Society of Biological Chemists conference, Philadelphia, PA, USA, 7 Jun 1987
Research Org:
Indiana Univ. School of Medicine, Indianapolis
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ENZYME INHIBITORS; BIOCHEMICAL REACTION KINETICS; OXIDOREDUCTASES; ENZYME ACTIVITY; PHOSPHORYLATION; AUTORADIOGRAPHY; ELECTROPHORESIS; PHOSPHATES; POTASSIUM COMPOUNDS; STEADY-STATE CONDITIONS; ALKALI METAL COMPOUNDS; CHEMICAL REACTIONS; ENZYMES; KINETICS; OXYGEN COMPOUNDS; PHOSPHORUS COMPOUNDS; REACTION KINETICS 550201* -- Biochemistry-- Tracer Techniques