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Purification of recombinant human pepsinogens and their application as immunoassay standards
Author(s) -
Aoki Takashi,
Tomaki Emiko,
Satoh Miyoko,
Takashiro Miho,
Onagi Hitoshi,
Itoh Masao,
Teramoto Tetsuya,
Morikawa Junji,
Watabe Hiroyuki
Publication year - 1998
Publication title -
iubmb life
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.132
H-Index - 113
eISSN - 1521-6551
pISSN - 1521-6543
DOI - 10.1080/15216549800202662
Subject(s) - maltose binding protein , immunoassay , recombinant dna , pepsin , escherichia coli , fusion protein , chemistry , biochemistry , microbiology and biotechnology , chromatography , biology , antibody , enzyme , immunology , gene
Human pepsinogen (PG) A and C were cloned in Escherichia coli, but the levels of expression were low and unstable. When these were fused to maltose‐binding protein (MBP), the fusion proteins (MBP‐PGA and MBP‐PGC) were expressed as the major products. Although these fused products were almost totally recovered from the insoluble fraction, the renaturation and purification procedures were easy and simple. MBP‐PGA and the PGA segment obtained by factor Xa digestion (designated as r‐PGA) possessed proteolytic activities equivalent to native PGA purified from gastric tissue (t‐PGA). For PGCs (MBP‐PGC, r‐PGC and t‐PGC) also, the specific activities were almost the same. However, the activities of PGCs were about 3‐ to 4‐hold higher than those of PGAs. In PGA and PGC immunoassay systems, r‐PGs (r‐PGA and r‐PGC) and the EIA kit standard PGs (gastric mucosal PGs) exhibited a good correlation. From these results, r‐PGs would seem to be applicable as assay standards without compromising the sensitivity of the immunoassay systems.