Functional roles of specific bruchid protease isoforms in adaptation to a soybean protease inhibitor.
Upon challenge by the soybean cysteine protease inhibitor soyacystatin N (scN), cowpea bruchids reconfigure their major digestive cysteine proteases (CmCPs) in adaptation to the inhibitor and resume normal feeding and development. We have previously shown that CmCPB transcripts were 116.3-fold more abundant in scN-adapted bruchid guts than in unadapted guts, while CmCPA transcripts were only 2.5-fold higher. In order to further elucidate the functional significance of this differential regulation, we expressed three CmCPA and one CmCPB isoforms (A9, A13, A16 and B1) using a bacterial expression system, and characterized their activities. In contrast to the precursors of CmCPAs (proCmCPAs), proCmCPB1 exhibited more efficient autocatalytic conversion from the latent proenzyme to its active mature protease form, and demonstrated higher intrinsic proteolytic activity. Among proCmCPAs, dependence on exogenous enzymatic processing varies: while maturation of proCmCPA13 and proCmCPA16 was impaired in the absence of external proteolytic activity, proCmCPA9 appeared to utilize a two-step autoprocessing mechanism. Although all CmCPs are scN-sensitive, scN was degraded by CmCPB1 when outnumbered by the protease, but scN remained intact in the presence of excessive CmCPA9. These results provide further evidence that differential expression of CmCPs under scN challenge brings about adaptation to the inhibitor. High induction of unique cysteine protease isoforms with superior autoprocessing and proteolytic efficacy represents a strategy cowpea bruchids use to cope with dietary scN.
Ahn
J-E
JE
Department of Entomology, Texas A & M University, College Station, TX 77843, USA.
Salzman
R A
RA
Braunagel
S C
SC
Koiwa
H
H
Zhu-Salzman
K
K
D000595:Amino Acid Sequence; D000818:Animals; D001517:Coleoptera / Q000201:enzymology*; D015891:Cystatins / Q000378:metabolism*; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D004064:Digestive System / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D015151:Immunoblotting; D007527:Isoenzymes; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D030262:Soybean Proteins
D015891:Cystatins; D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D030262:Soybean Proteins; C414109:scN protein, soybean; D010447:Peptide Hydrolases