The Primary Structure of the Heavy Chain of Chicken Gizzard Myosin

为了在分子水平上研究肌肉收缩，可能有必要揭示肌凝蛋白的初级结构，肌凝蛋白是收缩器官的主要组成部分。本研究测定了鸡胗肌球蛋白1重链S-1的n端203残基和c端203残基序列。在ATP存在下，用n -碘乙酰-n '-（5-磺基-1-萘基）乙二胺（简称IAEDANS）修饰Gizzard肌球蛋白。在5 M胍- hcl存在下，用Sephadex G-100柱进行凝胶过滤，在8 M尿素存在下，用CM 52柱进行层析，从修饰后的肌球蛋白的胰蛋白酶消化中分离出一个24千道尔的荧光片段。用常规方法测定该片段的氨基酸序列。该片段含有203个氨基酸残基和一个阻断的n端，被定位为重链的n端部分。在骨骼肌蛋白的同源位置发现了<epsilon> - n -三甲基赖氨酸，但没有<epsilon> - n -单甲基赖氨酸。兔骨肌球蛋白重链中被认为是atp结合残基之一的Trp-130被砂眼肌球蛋白中的谷氨酰胺所取代。该片段的Cys-93是与IAEDANS反应改变砂眼肌球蛋白atp酶活性的氨基酸残基。在没有ATP的情况下，用IAEDANS修饰砂眼肌球蛋白，用木瓜蛋白酶消化。从该摘要中分离出Mr=23,800的荧光片段，并按上述方法进行测序。木瓜蛋白酶片段含有203个氨基酸残基，与骨骼肌蛋白序列同源，可定位为S-1重链的c端。该片段的c端53个残基构成了肌球蛋白的颈部部分，含有13个带正电的残基，但没有带负电的残基。我们正在对位于上述两个片段之间的50千道尔顿片段进行测序。砂囊肌球蛋白头的初级结构在不久的将来可能会被揭示。少

It may be necessary to reveal the primary structure of myosin, a major component of the contractile apparatus, for studies of muscle contraction at the molecular level. In the present study, we determined the N-terminal 203-residue and C-terminal 203-residue sequences of the S-1 heavy chain of chicken gizzard myosin.1. Gizzard myosin was modified with N-iodoacetyl-N'-(5-sulfo-1-naphtyl)ethylenediamine (abbreviated as IAEDANS) in the presence of ATP. From the tryptic digest of the modified myosin, a fluorescent fragment (24 kilodalton) was isolated by gel filtration on a Sephadex G-100 column in the presence of 5 M guanidine-HCl followed by chromatography on a CM 52 column in the presence of 8 M urea. The amino acid sequence of the fragment was determined by conventional methods. The fragment contained 203 amino acid residues and a blocked N-terminus, and was assigned as an N-termnal part of the heavy chain. An <epsilon> -N-trimethyllysine was recognized at the homologous position with … More skeletal myosin, but no <epsilon> -N-monomethyllysine. Trp-130 of rabbit skeletal myosin heavy chain which is considered to be one of ATP-binding residue was replaced by gultamine in gizzard myosin. Cys-93 of the fragment was the amino acid residue whose reaction with IAEDANS alters the ATPase activity of gizzard myosin.2. Gizzard myosin was modified with IAEDANS in the absence of ATP, and digested with papain. From the digest, a fluorescent fragment of Mr=23,800 was isolated, and sequenced as described above. The papain-fragment contained 203 amino acid residues which could be assigned as C-terminal part of the S-1 heavy chain based on the homology with the sequence of skeletal myosin. The C-terminal 53 residues of this fragment, constituting the neck part of myosin, contained 13 positively charged residues but no negatively charged residue.3. We are sequencing the 50 kilodalton fragment which lies between the above two fragments. The primary structure of the gizzard myosin head could be revealed in the near future. Less