Rabbit plasma alpha-1-antiproteinase S-1: cloning, sequencing, expression, and proteinase inhibitory properties of recombinant protein.
A cDNA clone coding for the isoform S-1 of alpha-1-antiproteinase (also called alpha-1-proteinase inhibitor or alpha-1-antitrypsin) was isolated from rabbit liver cDNA library and sequenced. The cDNA consists of 1,426 nucleotides including 5' and 3' noncoding regions and codes for 413 amino acid residues including a signal peptide of 24 residues. The nucleotide and deduced amino acid sequences show 95.5 and 95.2% homologies, respectively, with the F isoform which occurs more abundantly in the rabbit serum than the S isoform. Of the 20 amino acid differences between the two isoforms, nine are located in a stretch of 15 amino acids encompassing the reactive site region, suggesting that these genes have diverged from each other by a nonrandom mechanism. A hypothesis is proposed that the domestication of animal is responsible for the extremely high evolutionary rate in the serpin reactive site region. Prokaryotic expression plasmids were constructed from the cDNA, transfected into Escherichia coli, and expressed. Partially purified recombinant protein inhibited elastase, but did not inhibit trypsin when a small substrate was used. The recombinant S-1 form, however, protected trypsin from inactivation by soybean trypsin inhibitor, a property characteristic of alpha-macroglobulins or rodent murinoglobulins. It is known that there are two types of interaction between serpin and proteinase: (i) most serpins form a stable equimolar complex with the enzyme, resulting in the enzyme inhibition and (ii) some serpins act as a substrate rather than as an inhibitor, resulting in the loss of inhibitory activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Biochemistry, Kinki University School of Medicine, Osaka.
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Saito
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Sinohara
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000032:analysis; D004926:Escherichia coli; D015870:Gene Expression / Q000235:genetics; D007536:Isomerism; D008969:Molecular Sequence Data; D011480:Protease Inhibitors; D011817:Rabbits; D011994:Recombinant Proteins; D017421:Sequence Analysis; D000515:alpha 1-Antitrypsin / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology*
D011480:Protease Inhibitors; D011994:Recombinant Proteins; D000515:alpha 1-Antitrypsin; D004247:DNA