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17. Isolation of a rat histidase cDNA sequence and expression and expression in Escherichia coli.

Author

Sano, Hirofumi, Tada, Toyohiro, Moriyama, Akihiko, Ogawa, Hisamitsu, Asai, Kiyofumi, Kawai, Yoko, Hodgson, Mark Emory, Kato, Taiji, Wada, Yoshiro, and Suchi, Mariko

Subjects
*PROTEINS, *EPITHELIUM, *IMMUNOGLOBULINS, *ESCHERICHIA coli, *GENETICS, *GENES
Abstract

Histidase (histidine ammonia-lyase) is a cytosolic enzyme responsible for catalyzing the non-oxidative deamination of histidine to urocanic acid. Full-length cDNAs encoding rat histidase have been isolated from a λZAP liver cDNA library using a partial cDNA fragment obtained by PCR. Whereas the initial description of the rat histidase 3′ untranslated sequence contained a rare polyadenylation signal sequence, the data presented encompass a more distant 28-bp region, possessing a nucleotide stretch (AATATAAA), identical to that in the mouse histidase cDNA. Dideoxynucleotide chain termination sequencing of two clones obtained by in vivo excision yielded an additional 376 bp and 105 bp of 5′ and 3′ untranslated sequences, respectively. A selected rat histidase cDNA clone was introduced into the pET-16b prokaryotic vector and expressed in BL21 (DE3)pLysS Escherichia coli. After purification by nickel-chelation chro- matography, recombinant histidine-tagged protein was employed to raise anti-(rat histidase) immunoglobulin in a Japanese white rabbit. The polyclonal rabbit antibody recognized and formed immune complexes with rat and recombinant human histidase proteins. Immunoblots of crude rat organ extracts detected a spectrum of histidase expression extending beyond that observed in liver and skin. Among other histidase-positive cells were those of the renal cortex tubular epithelium, fundic mucosal glands of stomach, gastric intramuscular (Auerbach's) plexus, and adrenal cortex. Immunohistochemical studies of histidase in rat liver produced discrete staining of hepatocytes in association with portal triads (Rappaport zone 1). Furthermore, in contrast with previous reports of activity confined to epidermal stratum corneum, our findings demonstrate immunoreactive protein within and limited to the adjacent stratum granulosum. [ABSTRACT FROM AUTHOR]

Published
1997
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