Amino Acid Sequence of Silkworm (Bombyx mori) Hemolymph Antitrypsin Deduced from Its cDNA Nucleotide Sequence: Confirmation of Its Homology with Serpins1

Abstract

A cDNA clone of silkworm (Bombyx mori) larval hemolymph antitrypsin (sw-AT) has been isolated from a fat body cDNA library. The cDNA has an open reading frame which codes a 392-amino acid residue polypeptide comprising a 16-residue signal peptide and a 376-residue mature sw-AT of Mr 41,805. The reactive site of sw-AT for inhibition of bovine trypsin [Sasaki, T. et al. (1987) J. Biochem. 102, 433-441] was identified as Lys343-Val344. Alignment of the sw-AT amino acid sequence with those of 11 members of the serpin superfamily of proteins clearly confirmed the homology of sw-AT with serpins. The amino acid sequence of sw-AT is 56% identical with that of the proteinase inhibitor from a lepidopteron, Manduca sexta [Kanost, M.R. et al. (1989) J. Biol. Chem. 264, 965-972], but the sequence around the reactive site shows no homology and the inhibitory specificity for proteinases is very different.

Differential expansion and evolution of the exon family encoding the Serpin-1 reactive centre loop has resulted in divergent serpin repertoires among the Lepidoptera

“…Mc Serpin-1j was similar to P. xylostella Serpin-1b, M. sexta Serpin-1A and B. mori AT. BmAT is an anti-trypsin with the P1-P1′ cleavage site residing between a Lys 343 -Val 344 (Takagi et al, 1990). Each serpin possessed a charged amino acid (Lys or Arg) at the analogous P1 site and were predicted to be trypsin inhibitors.…”

“…This causes the protease to relocate to the opposite pole of the serpin, renders the interaction irreversible, disorganizes the protease structure and increases its susceptibility to degradation (Huntington et al, 2000). Insect protease inhibitors resembling serpins were first isolated from the hemolymph of silkworm larvae, Bombyx mori, (Sasaki and Kobayashi, 1984) and later found to be closely related to a serpin from the tobacco hornworm, Manduca sexta (Kanost et al, 1989;Takagi et al, 1990). Additional insect Serpin genes have since been identified and much effort has been focused on their role in aspects of the insect defense response.…”

Codon usage patterns among genes for lepidopteran hemolymph proteins

“…Except as noted, sequences were obtained from GenBank. These were Manduca sexta: Ala-serpin, microvitellogenin ), apolipophorin III (Cole et al 1987), arylphorin alpha-and beta-subunits (Willot et al 1989, methionine-rich storage protein, transferfin, hemolin, cecropin B5P; Bombyx mori: 30K proteins 1-5 (Sakai et al 1988), antitrypsin (Takagi et al 1990), methionine-rich storage protein (Sakurai et al 1988), arylphorin (Fnjii et al 1989); Heliothis zea: juvenile hormone (JH) esterase; Hyalophora cecropia: cecropins A-D (Lidholm et al 1987;van Hofsten et al 1985), lysozyme ; and Trichoplusia ni: acidic-JH-suppressible protein 1 (Jones et al 1990). Simple tests of bias for a given base at the third position among the codons for a given amino acid were assessed by chi-square contingency.…”

Silk gland-specific proteinase inhibitor serpin16 from the Bombyx mori shows cysteine proteinase inhibitory activity

“…Among these serpins, only a few have been studied. Initially, two serpins, antichymotrypsin and antitrypsin, were identified in the hemolymph of B. mori [18,19]. In the 1990's, an additional antichymotrypsin was cloned from the larval fat body and termed antichymotrypsin II [20].…”