Phosphorylation of caldesmon prevents its interaction with smooth muscle myosin.
Phosphorylation of caldesmon prevents its interaction with smooth muscle myosin.
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发表时间：
1989-01
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影响因子：
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通讯作者：
C. Sutherland;M. Walsh
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作者：
C. Sutherland;M. Walsh
已知钙调素与平滑肌肌球蛋白结合。钙调素依赖的钙调素磷酸化完全阻断了钙调素与肌球蛋白的相互作用。2-硝基-5-硫氰基苯甲酸（NTCB）在其2个半胱氨酸残基处切割钙调蛋白，最初发生在一个位点，产生108-kDa和21.2-kDa肽，随后发生在108-kDa肽内的第二个位点，产生85-kDa和23.5-kDa片段。23.5-kDa肽保留了与肌球蛋白结合的能力。分离钙调蛋白的N-末端（95 kDa）和C-末端（42 kDa）胰凝乳蛋白酶肽，并用NTCB消化：C-末端肌动蛋白和钙调蛋白结合肽未被切割，表明其不含半胱氨酸残基，而95 kDa N-末端肽在两个位点被切割，产生56 kDa，23.5 kDa和21.2 kDa片段。因此，NTCB片段在caldesmon中的排列是：从N端到C端为21.2 kDa/23.5 kDa/85 kDa。用NTCB消化磷酸化的钙调蛋白，表明在21.2-kDa肽中有一个磷酸化位点，在23.5-kDa肽中有三个位点。这些结果导致了一个模型的发展，其中钙调蛋白可以交联肌动蛋白肌球蛋白和这种交联被钙调蛋白的磷酸化阻断。这一机制可以解释可逆的“闩桥”的形成，允许力维持在低水平的肌球蛋白磷酸化在完整的平滑肌。
Caldesmon is known to bind to smooth muscle myosin. Ca2+/calmodulin-dependent phosphorylation of caldesmon completely blocks its interaction with myosin. Cleavage of caldesmon at its 2 cysteine residues by 2-nitro-5-thiocyanobenzoic acid (NTCB) occurs initially at one site to yield 108-kDa and 21.2-kDa peptides and subsequently at the second site within the 108-kDa peptide to yield 85-kDa and 23.5-kDa fragments. The 23.5-kDa peptide retains the ability to bind to myosin. The N-terminal (95 kDa) and C-terminal (42 kDa) chymotryptic peptides of caldesmon were isolated and digested with NTCB: the C-terminal actin- and calmodulin-binding peptide was not cleaved, indicating that it does not contain either of the cysteine residues, whereas the 95-kDa N-terminal peptide was cleaved at two sites to yield 56-kDa, 23.5-kDa, and 21.2-kDa fragments. The arrangement of NTCB fragments in caldesmon is, therefore: 21.2 kDa/23.5 kDa/85 kDa from N to C terminus. Digestion of phosphorylated caldesmon with NTCB suggested a single phosphorylation site in the 21.2-kDa peptide and three sites in the 23.5-kDa peptide. These results lead to the development of a model whereby caldesmon may cross-link actin to myosin and such cross-linking is blocked by phosphorylation of caldesmon. This mechanism may explain the formation of reversible "latch bridges" which permit force maintenance at low levels of myosin phosphorylation in intact smooth muscles.