CALCIUM AND CALMODULIN IN SMOOTH MUSCLE CONTRACTION

钙调蛋白（Calmodulin, CaM）是一种普遍存在的钙结合蛋白

Calmodulin (CaM) is a ubiquitous Ca binding protein that has emerged as a universal regulator of the calcium signal. As cytosolic Ca rises, Ca binds to CaM producing structural changes which allow it to bind and activate more than thirty different target proteins. CaM is directly involved in the Ca dependent regulation of smooth muscle contraction, glandular and neurosecretion, DNA replication, gene expression and cell division. The present studies will utilize our new biologically active fluorescent CaM's which selectively report CaM binding to its target proteins. We will determine and relate the Ca dependence of CaM binding to its target proteins in smooth muscle to the Ca dependence of their activation. Using fluorescence stopped-flow techniques we will determine the rates of the Ca dependent association and dissociation of CaM with its target proteins, including myosin light chain kinase (MLCK) and caldesmon. These rates will be compared to the off-rates of calcium from CaM-target protein complexes. These studies will provide temporal information relating Ca transients (such as those muscle) with the rate of CaM-target protein association and disassociation. Most external stimuli influence intracellular events by the activation of cellular kinases and the phosphorylation of specific substrate proteins. Recently, it has been shown that several tyrosine kinases (including, the insulin receptor and the oncogene product, P60-src) are capable of phosphorylating tyrosine residues in Ca binding sites III and IV of CaM. We will examine the effect of this phosphorylation on CaM's calcium (and drug) binding properties and on the Ca dependence of CaM interaction with and activation of its target proteins. If phosphorylated CaM exhibition altered Ca dependent activation of MLCK, it will be examined for its ability to regulate Ca dependent tension in chemically skinned coronary arteries. Because CaM regulates many aspects of cell metabolism and growth its phosphorylation by these tyrosine kinases might play a pivotal role in the mechanism of action of Insulin and in the transformation of cells by oncogenic viruses which code for tyrosine kinases.