Muno D, Sekine T
Department of Biochemistry, School of Medicine, Juntendo University, Tokyo.
J Biochem. 1988 Sep;104(3):427-32. doi: 10.1093/oxfordjournals.jbchem.a122484.
We reported in the preceding paper [Muno, D., et al. (1987) J. Biochem. 101, 661-669] that the dinitrophenyl group exclusively introduced to SH1 on the 20-kDa fragment of myosin subfragment 1 was cross-linked to the 50-kDa fragment by irradiation, and that limited trypsinolysis of the cross-linked S1 generated an 83-kDa peptide, a cross-linking product between the 20- and 50-kDa fragments. This paper will deal with the location of the cross-linked residue on the 50-kDa fragment. When the 83-kDa fragment labeled at SH2 with a fluorogenic SH reagent was subjected to bromocyanolysis, a main fluorescent band, which implied a cross-linked peptide, appeared in the position with an apparent molecular mass of 18.5-kDa on SDS-PAGE. On the other hand, another cross-linked peptide was obtained from a complete tryptic digest of a 83-kDa fragment rich fraction. Amino acid sequence analysis of the two cross-linked peptides revealed that the DNP moiety attached at SH1 was cross-linked with a residue in the segment of the heavy chain spanning the 485-493 region from the N-terminus of the heavy chain.
我们在前一篇论文[Muno, D., 等人 (1987) J. Biochem. 101, 661 - 669]中报道，在肌球蛋白亚片段1的20-kDa片段上专门引入到SH1的二硝基苯基基团通过辐射与50-kDa片段交联，并且对交联的S1进行有限的胰蛋白酶消化产生了一个83-kDa的肽，这是20-kDa和50-kDa片段之间的交联产物。本文将探讨交联残基在50-kDa片段上的位置。当用荧光SH试剂在SH2处标记的83-kDa片段进行溴化氰裂解时，在SDS-PAGE上表观分子量为18.5-kDa的位置出现了一条主要的荧光带，这意味着是一个交联肽。另一方面，从富含83-kDa片段的完全胰蛋白酶消化物中获得了另一个交联肽。对这两个交联肽的氨基酸序列分析表明，连接在SH1上的DNP部分与重链中从N端起跨越485 - 493区域的片段中的一个残基交联。
