Green C R, Harfst E, Gourdie R G, Severs N J
Department of Anatomy and Embryology, University College London, U.K.
Proc R Soc Lond B Biol Sci. 1988 Mar 22;233(1271):165-74. doi: 10.1098/rspb.1988.0016.
The major gap junction polypeptide in most tissues has an apparent molecular mass of 27 kDa with a 47 kDa dimer present in junction-enriched fractions. However, a 54 kDa protein recognized by gap junction-specific antibodies has been reported and a complementary DNA (cDNA) sequence for both human and rat liver gap junctions codes for a 32 kDa protein. In this paper we show that these are all forms of the same gap junction protein that can be observed on SDS-polyacrylamide gels simply by varying the concentration of acrylamide in the gels. A 64 kDa dimer is also obtainable. Antibodies to the gap junction protein or to a synthetic peptide constructed to match the rat liver gap junction amino-terminal sequence recognize all of these forms. Under some conditions a 54 kDa dimer is 'preferred', explaining the presence of this species in whole tissue homogenate Western blots. These results clarify several controversies and indicate that the protein forming the gap junction channel probably undergoes no major post-translational modification as the cDNA sequence codes for a protein of molecular mass 32 kDa and this protein species and its 64 kDa dimer are demonstrable on SDS-polyacrylamide gels under appropriate conditions.
大多数组织中的主要间隙连接多肽的表观分子量为27 kDa，在富含连接的组分中存在47 kDa的二聚体。然而，已经报道了一种被间隙连接特异性抗体识别的54 kDa蛋白质，并且人和大鼠肝脏间隙连接的互补DNA（cDNA）序列编码一种32 kDa的蛋白质。在本文中，我们表明这些都是同一间隙连接蛋白的不同形式，只需改变凝胶中丙烯酰胺的浓度，就可以在SDS-聚丙烯酰胺凝胶上观察到。还可获得64 kDa的二聚体。针对间隙连接蛋白或为匹配大鼠肝脏间隙连接氨基末端序列而构建的合成肽的抗体可识别所有这些形式。在某些条件下，54 kDa的二聚体是“首选”的，这解释了该物种在全组织匀浆蛋白质免疫印迹中的存在。这些结果澄清了几个争议，并表明形成间隙连接通道的蛋白质可能没有经历重大的翻译后修饰，因为cDNA序列编码一种分子量为32 kDa的蛋白质，并且在适当条件下，这种蛋白质及其64 kDa的二聚体可在SDS-聚丙烯酰胺凝胶上显示出来。
