Partial purification and characterization of trypsin-like proteinases from insecticide-resistant and -susceptible strains of the maize weevil, Sitophilus zeamais

Comp Biochem Physiol B Biochem Mol Biol. 2010 Jan;155(1):12-9. doi: 10.1016/j.cbpb.2009.09.011. Epub 2009 Oct 14.


Serine proteinases from three strains of Sitophilus zeamais (Coleoptera: Curculionidae), one susceptible and two resistant to insecticides--one exhibiting fitness cost (resistant cost strain) and the other lacking it (resistant no-cost strain), were partially purified using an aprotinin-agarose affinity column providing purification factors ranging from 36.5 to 51.2%, with yields between 10 and 15% and activity between 529 and 875 microM/min/mg protein with the substrate N-alpha-benzoyl-L-Arg-p-nitroanilide (L-BApNA). SDS-PAGE of the purified fraction revealed a 56,000 Da molecular mass band in all strains and a 70,000 Da band more visible in the resistant no-cost strain. The purified proteinases from all strains were inhibited by phenylmethyl sulphonyl fluoride (PMSF), N-alpha-tosyl-L-lysine chloromethyl ketone (TLCK), aprotinin, benzamidine and soybean trypsin inhibitor (SBTI) characterizing them as trypsin-like serine proteinases. Trypsin-like proteinases from the resistant strains exhibited higher affinity for L-BApNA. The resistant no-cost strain exhibited V(max)-values 1.5- and 1.7-fold higher than the susceptible and resistance cost strains, respectively. A similar trend was also observed when using N-alpha-p-tosyl-L-Arg methyl ester (L-TAME) as substrate. These results provide support to the hypothesis that the enhanced serine proteinase activity may be playing a role in mitigating physiological costs associated with the maintenance of insecticide resistance mechanisms in some maize weevil strains.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Benzoylarginine Nitroanilide / metabolism
  • Catalysis / drug effects
  • Coleoptera / classification
  • Coleoptera / enzymology*
  • Electrophoresis, Polyacrylamide Gel
  • Hydrogen-Ion Concentration
  • Insect Proteins / isolation & purification
  • Insect Proteins / metabolism*
  • Insecticide Resistance*
  • Insecticides / toxicity*
  • Kinetics
  • Phenylmethylsulfonyl Fluoride / pharmacology
  • Protease Inhibitors / pharmacology
  • Serine Proteases / isolation & purification
  • Serine Proteases / metabolism*
  • Species Specificity
  • Substrate Specificity
  • Temperature
  • Trypsin / isolation & purification
  • Trypsin / metabolism
  • Zea mays / parasitology*


  • Insect Proteins
  • Insecticides
  • Protease Inhibitors
  • Phenylmethylsulfonyl Fluoride
  • Benzoylarginine Nitroanilide
  • Serine Proteases
  • Trypsin