Proteolytic activity in the extracts and in the excretory/secretory products from Varroa destructor parasitic mite of honeybee

Abstract

Proteolytic activity of the extracts and excretory–secretory (E/S) products from parasitic mite Varroa destructor of honeybees was determined and partially characterized. Proteolytic activity of mite extract was the highest at pH 3.5; the second smaller peak was at pH 5.0. After sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) 23 protein fractions (4.3–210 kDa) in mite extract and 15 protein fractions in E/S products were revealed. Three of them (50, 52 and 54 kDa) in mite extract and two (50 and 54 kDa) in E/S products were active towards albumin and gelatin. Peptidases from E/S products had higher activity (per 1 mg protein) than enzymes from whole mite extract. They more efficiently hydrolysed albumin and haemoglobin than gelatin and casein. Peptidases from both materials were sensitive to aspartyl and cysteine proteinase inhibitors (peptidase inhibitors, or PIs). Pepstatin A (aspartyl PI) acted the strongest and diminished the activity at pH 3.5 by 76.5% and by 86.8% at pH 5, l-trans-epoxysuccinyl-leucylamido-(4-guanidino)-butane (broad cysteine PI) by 20.1 and 33.7%, respectively. Proteolysis was slightly reduced only at pH 5 by ethylene diamine tetraacetic acid (metalloproteinase inhibitor) and serine peptidases, Kunitz and soybean trypsin inhibitors. These results suggested that the enzymes of V. destructor belong mainly to aspartyl and cysteine peptidases.