Burnham D B
Biochem J. 1985 Oct 15;231(2):335-41. doi: 10.1042/bj2310335.
Ca2+-activated protein phosphatase activity was demonstrated in mouse pancreatic acinar cytosol with alpha-casein and skeletal-muscle phosphorylase kinase as substrates. This phosphatase activity preferentially dephosphorylated the alpha subunit of phosphorylase kinase. After DEAE-cellulose chromatography, the Ca2+-activated phosphatase activity became dependent on exogenous calmodulin for maximal activity. Half-maximal activation was achieved at 0.5 +/- 0.1 microM-Ca2+. Trifluoperazine completely inhibited Ca2+-activated phosphatase activity, with half-maximal inhibition occurring at 8.5 +/- 0.6 microM. Mn2+, but not Mg2+, at 1 mM concentration could substitute for Ca2+ in eliciting full enzyme activation. The apparent Mr of the phosphatase as determined by Sephadex G-150 chromatography was 93000 +/- 1000. Submitting active fractions obtained after Sephadex chromatography to calmodulin affinity chromatography resulted in the resolution of a major protein of Mr 55500 +/- 300. In conclusion, Ca2+-activated protein phosphatase activity has been identified in exocrine pancreas and has several features in common with Ca2+-activated calmodulin-dependent protein phosphatases previously isolated from brain and skeletal muscle. It is possible that this Ca2+-activated phosphatase may utilize as substrates certain acinar-cell phosphoproteins previously shown to undergo dephosphorylation in response to Ca2+-mediated secretagogues.
以α-酪蛋白和骨骼肌磷酸化酶激酶为底物，在小鼠胰腺腺泡细胞溶质中证实了Ca2+激活的蛋白磷酸酶活性。这种磷酸酶活性优先使磷酸化酶激酶的α亚基去磷酸化。经DEAE-纤维素层析后，Ca2+激活的磷酸酶活性依赖于外源性钙调蛋白才能达到最大活性。在0.5±0.1μM-Ca2+时达到最大激活的一半。三氟拉嗪完全抑制Ca2+激活的磷酸酶活性，在8.5±0.6μM时出现最大抑制的一半。1 mM浓度的Mn2+而非Mg2+可替代Ca2+引发酶的完全激活。通过Sephadex G-150层析测定，该磷酸酶的表观相对分子质量为93000±1000。将Sephadex层析后获得的活性组分进行钙调蛋白亲和层析，得到一种主要蛋白质，其相对分子质量为55500±300。总之，在胰腺外分泌腺中已鉴定出Ca2+激活的蛋白磷酸酶活性，它与先前从脑和骨骼肌中分离出的Ca2+激活的钙调蛋白依赖性蛋白磷酸酶有几个共同特征。这种Ca2+激活的磷酸酶有可能将某些先前显示在Ca2+介导的促分泌剂作用下会发生去磷酸化的腺泡细胞磷蛋白作为底物。
