Baker J E, Fabrick J A
USDA, ARS, Grain Marketing and Production Research Center, 1515 College Avenue, KS 66502, Manhattan, USA.
Insect Biochem Mol Biol. 2000 Oct;30(10):937-46. doi: 10.1016/s0965-1748(00)00066-7.
Host plasma proteins and protein digestion in larval parasitoids were studied during trophic interactions of the ectoparasitoid Habrobracon hebetor Say (Hymenoptera: Braconidae), with a host, larvae of the Indianmeal moth, Plodia interpunctella Hübner (Lepidoptera: Pyralidae). We could detect no apparent differences in host hemolymph protein patterns up to 72 h after paralysation and/or parasitization by H. hebetor. A 190 kDa putative apolipophorin I present in host hemolymph could not be detected in the midguts of feeding H. hebetor larvae indicating that it is rapidly digested. The major 60 kDa storage proteins (putative hexamerins) in host hemolymph were detected in the parasitoid midgut and were completely digested 24 h after cessation of feeding and the beginning of cocoon formation. Host hemolymph had a pH of about 6.4. The pH optima of the midgut proteinases in the larval parasitoid were in the alkaline region, but midgut fluid in feeding parasitoid larvae was about pH 6. 8. Based on enzyme activity against selected artificial proteinase substrates including azocasein, N-alpha-benzoyl-L-Arg p-nitroanilide (BApNA), succinyl-Ala-Ala-Pro-Phe p-nitroanilide (SAAPFpNA), succinyl-Ala-Ala-Pro-Leu p-nitroanilide (SAAPLpNA), and inhibition by selected proteinase inhibitors, serine proteinases appear to be the predominant class of enzymes involved in protein digestion in the midguts of H. hebetor. There is also an active aminopeptidase (LpNA) associated with the microsomal fraction of midgut preparations. There was no evidence for preoral digestion or ingestion of proteinases from host hemolymph by the parasitoid larva. There was a very active BApNAase in the soluble fraction of midgut extracts. This activity increased on a per midgut basis up to 24 h after the beginning of cocoon formation but decreased rapidly by 48 h. Two major (P1 and P3) and several minor proteinases were detected in midgut extracts of H. hebetor analysed with gelatin zymograms. The apparent molecular mass of P1 varied from 95 to 49 kDa depending on protein loading. P3 had an apparent molecular mass of 39 kDa that was independent of protein loading. In summary, electrophoretic evidence indicates that host hemolymph protein patterns do not change significantly for at least 72 h after paralysation by H. hebetor. The role, if any, of envenomization in preventing breakdown of hemolymph proteins during this time remains to be determined. Because the predominant host hemolymph proteins, a putative apolipophorin I and the putative hexamerins, are readily digested by the serine proteinases present in the midguts of this parasitoid larva, these or similar proteins would provide an easily digested source of dietary amino acids that could be used for development of artificial diets for this beneficial insect.
在体外寄生蜂哈氏肿腿蜂（Habrobracon hebetor Say，膜翅目：茧蜂科）与寄主印度谷螟（Plodia interpunctella Hübner，鳞翅目：螟蛾科）幼虫的营养相互作用过程中，研究了寄主血浆蛋白以及寄生蜂幼虫体内的蛋白消化情况。在哈氏肿腿蜂使寄主麻痹和/或寄生后的72小时内，我们未检测到寄主血淋巴蛋白模式有明显差异。寄主血淋巴中存在的一种190 kDa的假定载脂蛋白I，在取食的哈氏肿腿蜂幼虫中肠中未被检测到，这表明它被迅速消化。寄主血淋巴中的主要60 kDa储存蛋白（假定六聚蛋白）在寄生蜂中肠中被检测到，并在停止取食和开始结茧24小时后被完全消化。寄主血淋巴的pH约为6.4。寄生蜂幼虫中肠蛋白酶的最适pH在碱性区域，但取食的寄生蜂幼虫中肠液的pH约为6.8。基于对包括偶氮酪蛋白、N-α-苯甲酰-L-精氨酸对硝基苯胺（BApNA）、琥珀酰-Ala-Ala-Pro-Phe对硝基苯胺（SAAPFpNA）、琥珀酰-Ala-Ala-Pro-Leu对硝基苯胺（SAAPLpNA）在内的选定人工蛋白酶底物的酶活性，以及选定蛋白酶抑制剂的抑制作用，丝氨酸蛋白酶似乎是参与哈氏肿腿蜂中肠蛋白消化的主要酶类。中肠制剂的微粒体部分还存在一种活性氨肽酶（LpNA）。没有证据表明寄生蜂幼虫存在口前消化或摄取寄主血淋巴中的蛋白酶。中肠提取物的可溶部分有非常活跃的BApNA酶活性。这种活性在结茧开始后每中肠的基础上在24小时内增加，但在48小时时迅速下降。用明胶酶谱分析哈氏肿腿蜂的中肠提取物时，检测到两种主要蛋白酶（P1和P3）和几种次要蛋白酶。P1的表观分子量根据蛋白上样量在95至49 kDa之间变化。P3的表观分子量为39 kDa，与蛋白上样量无关。总之，电泳证据表明，在哈氏肿腿蜂使寄主麻痹后至少７２小时内，寄主血淋巴蛋白模式没有显著变化。在此期间，若毒液对防止血淋巴蛋白分解有作用，其作用仍有待确定。由于寄主血淋巴中的主要蛋白，即假定的载脂蛋白I和假定的六聚蛋白，很容易被这种寄生蜂幼虫中肠中的丝氨酸蛋白酶消化，这些蛋白或类似蛋白将提供一种易于消化的膳食氨基酸来源，可用于为这种有益昆虫开发人工饲料。
