Activation of myosin light chain kinase and nitric oxide synthase activities by calmodulin fragments.

作者：Anthony Persechini, Kirk McMillan, P LEAKEY · 发表于：Journal of Biological Chemistry · 年份：1994 · DOI：10.1016/s0021-9258(17)33985-6 · 被引用次数：86 · 研究领域：Protein Kinase Regulation and GTPase Signaling、Mitochondrial Function and Pathology、ATP Synthase and ATPases Research

We have investigated the abilities of calmodulin (CaM) tryptic fragments 1-75 (TRCI) or 78-148 (TRCII) to activate gizzard smooth muscle myosin light chain kinase (gMLCK), rabbit skeletal muscle myosin light chain kinase (skMLCK), and neural nitric oxide synthase (nNOS) activities. Our results indicate for all three enzymes that binding of CaM follows an ordered mechanism wherein the C-terminal lobe, represented by TRCII, binds specifically to a site we designated as A, followed by binding of the N-terminal lobe, represented by TRCI, to a site designated as B. With TRCII and TRCI bound to their respective sites, skMLCK and gMLCK activities are both activated to about 80% of their maximum levels. Occupancy of both sites in the MLCK enzymes by TRCI results in only low levels of enzyme activation; occupancy of both sites by TRCII also results in low levels of gMLCK activity, but activates skMLCK activity to 65% of the maximum level. With TRCI bound at site B and either TRCII or TRCI bound at site A, nNOS activity is 50% of the maximum level. Apparent dissociation constants for TRCII binding to site A and TRCI binding to site B are, respectively; 0.3 and 3 microM (skMLCK); 1.2 and 0.8 microM (gMLCK); 10 nM and 150 microM (nNOS). Our results demonstrate that the CaM lobes can make distinct contributions to binding and/or activation of different CaM-dependent enzymes and that the tethering function of the central helix can be mimicked by sufficiently high concentrations of the CaM ...