Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.1 (chymotrypsin)
 10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chymotryptic cleavage of the alpha-subunit of the canine kidney Na+/K(+)-ATPase in the presence of Na+ abolishes ATPase activity and yields an 83 kDa peptide from Ala 267 to the COOH-terminus. To test the proposal that E1 to E2 conformational transition is blocked in this modified enzyme, we have made a detailed comparison of its phosphorylation with that of the native enzyme by ATP. While phosphorylation of alpha is dependent on Na+ and prevented by K+, that of the 83 kDa peptide is modestly stimulated by Na+; and only this stimulation, but not the Na(+)-independent phosphorylation is inhibited by K+. Ouabain, which inhibits alpha-phosphorylation by ATP, activates Na(+)-independent phosphorylation of the 83 kDa peptide by ATP, and inhibits the Na(+)-stimulation of this process. While there is a ouabain-stimulated phosphorylation of alpha by Pi, the 83 kDa peptide is not phosphorylated by Pi with or without ouabain. In its sensitivity to ADP, and insensitivity to K+, the phosphopeptide is similar to the E1P of the native enzyme; however, the spontaneous decomposition rate of the phosphopeptide is orders of magnitude lower than that of the native EP. Na+ has no effect on the spontaneous decomposition of the phosphopeptide; but at high Na+ concentrations (K0.5 = 350 mM) the ADP sensitivity of the phosphopeptide is reduced. The phosphopeptide, like the native EP, is acid-stable, alkaline-labile, and sensitive to hydroxylamine and molybdate. The chymotrypsin-treated enzyme catalyzes an ADP-ATP exchange activity that is stimulated by Na+. The Na(+)-independent part of this exchange, unlike that of the native enzyme, is activated by ouabain. Our findings establish that (a) the phosphorylation process and its control by Na+, K+ and ouabain are autoregulated by the NH2-terminal domain of the alpha-subunit; and (b) the often repeated assumption that the primary role of this domain is in the regulation of E1-E2 transitions is not valid.
 ...
PMID:Autoregulation of the phosphointermediate of Na+/K(+)-ATPase by the amino-terminal domain of the alpha-subunit. 217 3

1. Using a previously established method of isolating an active-sodium-transport inhibitor (ASTI) from hypothalamic cell culture medium, the inhibitor was isolated and partially purified from sequential passages through Sephadex G-25 and h.p.l.c., and its effects on de-endothelialized rabbit aortic strips were investigated. 2. ASTI caused a cumulative concentration-dependent increase in tension which reversed slowly after wash, and the wash showed an identical effect on fresh strips. 3. Ouabain, used as a control, also caused a concentration-dependent increase in tension which reached a plateau at a concentration of 10 mmol/l. Both ouabain and ASTI caused a significant potentiation of the vasoconstrictor effect of noradrenaline at concentrations of 1 nmol/l-0.1 mmol/l. 4. Both ASTI and ouabain caused a significantly greater (P less than 0.01) calcium retention than control medium in aortic strips. 5. Incubation of ASTI with prolidase, chymotrypsin and carboxypeptidase A destroyed the vasoconstrictor effects as well as its inhibitory effects on sodium, potassium-dependent adenosine triphosphatase and sodium efflux from erythrocytes, but leucine aminopeptidase was ineffective. 6. These studies suggest that hypothalamic cells in culture release a peptidic inhibitor of active sodium transport which increases vascular reactivity, potentiates vasoconstrictor effects of noradrenaline and causes calcium retention.
 ...
PMID:Calcium retention and increased vascular reactivity caused by a hypothalamic sodium transport inhibitor. 340 35

This study describes the biochemical composition of junctional feet in skeletal muscle utilizing a fraction of isolated triad junctions. [3H]Ouabain entrapment was employed as a specific marker for T-tubules. The integrity of the triad junction was assayed by the isopycnic density of [3H]ouabain activity (24-30% sucrose for free T-tubules, 38-42% sucrose for intact triads). Trypsin, chymotrypsin, and pronase all caused separation of T-tubules from terminal cisternae, indicating that the junction is composed as least in part of protein. Trypsin and chymotrypsin hydrolyzed four proteins: the Ca2+ pump, a doublet 325,000, 300,000, and an 80,000 Mr protein. T-tubules which had been labeled covalently with 125I were joined to unlabeled terminal cisternae by treatment with K cacodylate. The reformed triads were separated from free T-tubules and then severed by passage through a French press. When terminal cisternae were separated from T-tubules, some 125I label was transferred from the labeled T-tubules to the unlabeled terminal cisternae. Gel electrophoresis showed that, although T-tubules were originally labeled in a large number of different proteins, only a single protein doublet was significantly labeled in the originally unlabeled terminal cisternae. This protein pair had molecular weights of 325,000 and 300,000 daltons. Transfer of label did not occur to a substantial degree without K cacodylate treatment. We propose that the transfer of 125I label from T-tubules to terminal cisternae during reformation and breakage of the triad junction is a property of the protein which spans the gap between T-tubules and terminal cisternae.
 ...
PMID:Identification of a constituent of the junctional feet linking terminal cisternae to transverse tubules in skeletal muscle. 674 61