人前列腺特异性腺激肽释放酶hK2在大肠杆菌中的过表达及抗体的产生。

Overexpression of a human prostate-specific glandular kallikrein, hK2, in E. coli and generation of antibodies.

作者信息

Saedi M S, Cass M M, Goel A S, Grauer L, Hogen K L, Okaneya T, Griffin B Y, Klee G G, Young C Y, Tindall D J

机构信息

Hybritech Incorporated, San Diego, CA 92196, USA.

出版信息

Mol Cell Endocrinol. 1995 Apr 1;109(2):237-41. doi: 10.1016/0303-7207(95)03508-5.

DOI:10.1016/0303-7207(95)03508-5

PMID:7664987

Abstract

The genomic and the cDNA clones of human glandular kallikrein (hK2), a member of the kallikrein family, have been isolated; however, the hK2 protein has not yet been identified and characterized. The deduced sequence of hK2 is highly homologous to prostate specific antigen (PSA), a widely accepted prognostic indicator of prostate carcinoma. Also, hK2 mRNA, like PSA mRNA, is exclusively expressed in prostatic epithelia. These two properties make hK2 a potentially useful marker for studying prostate cancer. In this paper, we describe for the first time the overexpression of the entire hK2 protein (pre-pro hK2:pphK2) in the E. coli system. Our system yields high levels of authentic pphK2 (as determined by partial amino acid sequence analysis) comprising about 40% of total cellular protein. pphK2 was purified to near homogeneity by preparative SDS/PAGE and used to generate anti-pphK2 antibodies in rabbits. The antibodies recognize the recombinant hK2 protein and a major band of approximately 34 kDa in seminal fluid.

摘要

人腺体激肽释放酶（hK2）作为激肽释放酶家族的一员，其基因组克隆和cDNA克隆已被分离出来；然而，hK2蛋白尚未得到鉴定和表征。hK2的推导序列与前列腺特异性抗原（PSA）高度同源，PSA是前列腺癌广泛认可的预后指标。此外，hK2 mRNA与PSA mRNA一样，仅在前列腺上皮中表达。这两个特性使hK2成为研究前列腺癌的潜在有用标志物。在本文中，我们首次描述了在大肠杆菌系统中全长hK2蛋白（前原hK2：pphK2）的过表达。我们的系统产生了高水平的真实pphK2（通过部分氨基酸序列分析确定），其占细胞总蛋白的约40%。通过制备性SDS/PAGE将pphK2纯化至接近均一，并用于在兔体内产生抗pphK2抗体。这些抗体识别重组hK2蛋白以及精液中一条约34 kDa的主要条带。