The subtilisins of the invertebrate mycopathogens Verticillium chlamydosporium and Metarhizium anisopliae. are serologically and functionally related

Abstract

The major extracellular proteases from the nematophagous fungus Verticillium chlamydosporium and the entomophagous fungus Metarhizium anisopliae, VCP1 and Pr1, respectively, are closely related both functionally and serologically. Antibodies raised against either enzyme cross-reacted with both antigens, suggesting that they have common epitopes. The VCP1 and Pr1 antisera labelled bovine pancreatic elastase and proteinase K, respectively. Neither antiserum reacted with commercial chymotrypsin. An antiserum to a serine protease from the closely related V. suchlasporium also cross-reacted with VCP1 and Pr1. In contrast, a polyclonal antibody to an isoform of Pr1 exclusive to M. anisopliae isolate ME1 failed to recognize Pr1 from M. anisopliae V245 or VCP1. The N-terminal amino acid sequence of VCP1 revealed similarities with subtilisin-like enzymes from other fungi, but the closest match was with Pr1. The pure enzymes, VCP1 and Pr1, failed to hydrolyse mono-aminoacyl-naphthylamide substrates but demonstrated dipeptidyl peptidase activity against Gly-Pro-beta NA and Leu-Ala-beta NA, respectively. These results are discussed in the context of specificity of invertebrate mycopathogens.

Infection of plant-parasitic nematodes by nematophagous fungi – a review of the application of molecular biology to understand infection processes and to improve biological control

“…The fungus may also be able to adjust the pH of microenvironments to facilitate optimal enzyme activity. This was shown for the insect parasite Metarhizium anisopliae (St Leger et al, 1999) and may be the same for nematophagous egg parasites since their biology is similar (Segers et al, 1995). Adjustment of pH is important because suboptimal pH can suppress protease expression (St Leger et al, 1998).…”

“…Small differences in amino acid sequence can affect substrate utilisation, kinetics, or surface properties of the enzyme (Segers et al, 1995). The differences in substrate utilisation between Pr1 and VCP1 may be due to amino acid polymorphisms (Segers et al, 1995) because these are known to have profound effects on subtilases (Wells & Estell, 1988).…”

Expression of serine proteases in egg-parasitic nematophagous fungi during barley root colonization

“…Most of these proteolytic enzymes belong to the family of serine proteases (Lopez-Llorca et al, 2008). Specific polyclonal antibodies against the major extracellular proteases VCP1 and P32 from P. chlamydosporia and P. rubescens, respectively, have been obtained and shown cross-reactivity toward both proteins (Lopez-Llorca and Robertson, 1992;Segers et al, 1995). Polyclonal antibodies are also suitable for antigen immunolocalization purposes.…”

Characterizing structural features of cuticle-degrading proteases from fungi by molecular modeling

“…However, no polymorphism has been found at the catalytic residue triad of the Asp, His and Ser, indicating a strong selective pressure for the maintenance of the structure/function of this site. Some of the polymorphisms observed near the substrate-binding regions may have effects on the substrate utilization, kinetics or surface property of the enzymes [37]. It has been suggested that the abundant polymorphisms within PR1 may render its isoforms recalcitrant to some of the antiproteolytic hemolymphal peptides and could enable the pathogenic fungus M. anisopliae to exploit a wider range of substrates [38].…”