Purification and partial characterization of thermal hysteresis proteins from overwintering larvae of pine needle gall midge, Thecodiplosis japonensis (Diptera : Cecidomiidae)

摘要

The pine needle gall midge of Thecodiplosis japonensis is a serious forest pest and overwinters as a 3rd instar larva at soil surface in Korea. The time necessary for killing 50% of larvae at -15 degrees C is 160 min. During overwintering period, I: japonensis larvae accumulate relatively high content of trehalose as the main cryoprotectant. In this paper, the proteinaceous cryoprotectants were identified. Two thermal hysteresis proteins (THP-1S and 2S) were purified from overwintering larvae by ethanol fractionation, trichloroacetic acid precipitation, ion-exchange chromatography (DEAE-Sephadex A-25) and gel permeation chromatography (Sephadex G-100). Their molecular weights are 34.9 and 37.8 kD respectively. I: japonensis THPs cannot be stained by periodic acid-Schiffs' reagent, suggesting no carbohydrate in them. The thermal hysteresis activity of THP-2 at the concentration of 50 mg/ml is 11.02+/-0.08 degrees C (mean+/-SD, n=10), perhaps the highest active insect THP. It is the first report of purified II: japonensis THPs in Diptera. [References: 30]