Taggart R T, Cass L G, Mohandas T K, Derby P, Barr P J, Pals G, Bell G I
Department of Molecular Biology and Genetics, Wayne State University School of Medicine, Detroit, Michigan 48201.
J Biol Chem. 1989 Jan 5;264(1):375-9.
The entire pepsinogen C (PGC) coding sequence was determined by analysis of a series of five overlapping cDNA clones identified in a library constructed from human gastric mucosa poly(A+) RNA. A partial cDNA clone was initially identified using a 256-fold degenerate oligonucleotide probe for amino acid residues 4-12 of pepsin C, and subsequently 4 additional clones were identified upon rescreening with a probe complementary to the 5' region of the original cDNA clone. Northern analysis of gastric mucosa poly(A+) RNA with a PGC cDNA probe revealed an mRNA 1.5-kilobase species that was indistinguishable from that detected with a human pepsinogen A (PGA) cDNA probe. In contrast, the PGC and PGA cDNA probes detected distinct genomic restriction fragments indicating there was no detectable cross-hybridization under high stringency conditions. The PGC gene was localized to human chromosome 6 by analysis of a panel of human x mouse somatic cell hybrids. The regions containing the active site aspartyl groups of PGC are conserved in relationship to several other aspartic proteinases. We propose that the absence of detectable immunologic cross-reactivity between the two groups of human pepsinogens, A and C, results from divergent evolution of sequences located on the surface of the zymogens in contrast to the strongly conserved active site regions located within the binding cleft of the enzymes that are inaccessible for antigenic recognition.
通过分析从人胃黏膜多聚腺苷酸（poly(A+)）RNA构建的文库中鉴定出的一系列五个重叠cDNA克隆，确定了整个胃蛋白酶原C（PGC）编码序列。最初使用针对胃蛋白酶C第4至12个氨基酸残基的256倍简并寡核苷酸探针鉴定了一个部分cDNA克隆，随后用与原始cDNA克隆5'区域互补的探针重新筛选时又鉴定出另外4个克隆。用PGC cDNA探针对胃黏膜多聚腺苷酸（poly(A+)）RNA进行Northern分析，发现一种1.5千碱基的mRNA，与用人胃蛋白酶原A（PGA）cDNA探针检测到的mRNA无法区分。相比之下，PGC和PGA cDNA探针检测到不同的基因组限制性片段，表明在高严格条件下没有可检测到的交叉杂交。通过分析一组人×小鼠体细胞杂种，将PGC基因定位到人类6号染色体。与其他几种天冬氨酸蛋白酶相比，PGC含有活性位点天冬氨酰基团的区域是保守的。我们提出，两组人胃蛋白酶原A和C之间缺乏可检测到的免疫交叉反应性，是由于酶原表面序列的趋异进化，而位于酶结合裂隙内的高度保守的活性位点区域则无法进行抗原识别。
