Adam L P, Haeberle J R, Hathaway D R
Department of Medicine, Indiana University School of Medicine, Indianapolis 46202-4800, USA.
Am J Physiol. 1995 Apr;268(4 Pt 1):C903-9. doi: 10.1152/ajpcell.1995.268.4.C903.
Calponin and caldesmon were purified from porcine carotid arteries that were preincubated with [32P]orthophosphate, and the stoichiometry of phosphorylation was measured. In resting arteries, caldesmon was phosphorylated to a level of 0.41 mol PO4/mol protein, while calponin was phosphorylated to levels < 0.01 mol PO4/mol protein. Stimulation by histamine (1 or 5 min), KCl (1, 5 or 60 min), or phorbol 12,13-dibutyrate (PDBu; 1 microM for 15 or 60 min) did not lead to measurable increases in the PO4 content of calponin. Because dephosphorylation of calponin during the purification procedure could account for these results, we also determined stoichiometries after firat denaturing endogenous phosphatases with trichloroacetic acid. In these experiments, calponin was determined to be phosphorylated to the same low levels as in the first set of experiments. Collectively, these data show that calponin is not phosphorylated to significant levels during contractions of carotid arteries under conditions where caldesmon phosphorylation is apparent. The circumstances under which calponin may be phosphorylated in intact smooth muscle, and the purpose that may be served by this potential regulatory process, remain to be determined.
从预先用[32P]正磷酸盐孵育的猪颈动脉中纯化钙调蛋白和钙调素，并测量磷酸化的化学计量。在静息动脉中，钙调素的磷酸化水平为0.41摩尔磷酸根/摩尔蛋白质，而钙调蛋白的磷酸化水平<0.01摩尔磷酸根/摩尔蛋白质。组胺（1或5分钟）、氯化钾（1、5或60分钟）或佛波醇12,13-二丁酸酯（PDBu；1微摩尔，15或60分钟）刺激并未导致钙调蛋白的磷酸根含量出现可测量的增加。由于纯化过程中钙调蛋白的去磷酸化可能解释这些结果，我们还在用三氯乙酸使内源性磷酸酶变性后测定了化学计量。在这些实验中，钙调蛋白的磷酸化水平与第一组实验中相同，处于较低水平。总体而言，这些数据表明，在钙调素磷酸化明显的情况下，颈动脉收缩过程中钙调蛋白不会被磷酸化到显著水平。钙调蛋白在完整平滑肌中可能被磷酸化的情况以及这种潜在调节过程可能发挥的作用仍有待确定。
