Elvin C M, Vuocolo T, Pearson R D, East I J, Riding G A, Eisemann C H, Tellam R L
CSIRO Division of Tropical Animal Production, CSIRO Private Mail Bag 3, Indooroopilly, 4068, Queensland, Australia.
J Biol Chem. 1996 Apr 12;271(15):8925-35. doi: 10.1074/jbc.271.15.8925.
The peritrophic membrane is a semi-permeable chitinous matrix lining the gut of most insects and is thought to have important roles in the maintenance of insect gut structure, facilitation of digestion, and protection from invasion by microrganisms and parasites. Proteins are integral components of this matrix, although the structures and functions of these proteins have not been characterized in any detail. The peritrophic membrane from the larvae of the fly Lucilia cuprina, the primary agent of cutaneous myiasis in sheep, was shown to contain six major integral peritrophic membrane proteins. Two of these proteins, a 44-kDa glycoprotein (peritrophin-44) and a 48-kDa protein (peritrophin-48) together represent >70% of the total mass of the integral peritrophic membrane proteins. Peritrophin-44 was purified and its complete amino acid sequence was determined by cloning and sequencing the DNA complementary to its mRNA. The deduced amino acid sequence codes for a protein of 356 amino acids containing an amino-terminal signal sequence followed by five similar but nonidentical domains, each of approximately 70 amino acids and characterized by a specific register of 6 cysteines. One of these domains was also present in the noncatalytic regions of chitinases from Brugia malayi, Manduca sexta, and Chelonus. Peritrophin-44 has a uniform distribution throughout the larval peritrophic membrane. Reverse transcriptase-polymerase chain reaction detected the expression of peritrophin-44 in all three larval instars but only trace levels in adult L. cuprina. The protein binds specifically to tri-N-acetyl chitotriose and reacetylated chitosan in vitro. It is concluded that the multiple cysteine-rich domains in peritrophin-44 are responsible for binding to chitin, the major constituent of peritrophic membrane. Peritrophin-44 probably has roles in the maintenance of peritrophic membrane structure and in the determination of the porosity of the peritrophic membrane. This report represents the first characterization of an insect peritrophic membrane protein.
围食膜是一种半透性的几丁质基质，衬于大多数昆虫的肠道内，被认为在维持昆虫肠道结构、促进消化以及保护免受微生物和寄生虫入侵方面发挥着重要作用。蛋白质是这种基质的重要组成部分，尽管这些蛋白质的结构和功能尚未得到详细表征。羊皮肤蝇蛆病的主要病原体——绿蝇（Lucilia cuprina）幼虫的围食膜被证明含有六种主要的围食膜整合蛋白。其中两种蛋白质，一种44 kDa的糖蛋白（围食蛋白-44）和一种48 kDa的蛋白质（围食蛋白-48），它们共同占围食膜整合蛋白总质量的70%以上。围食蛋白-44被纯化，其完整的氨基酸序列通过克隆和测序与其mRNA互补的DNA来确定。推导的氨基酸序列编码一个由356个氨基酸组成的蛋白质，该蛋白质含有一个氨基末端信号序列，随后是五个相似但不相同的结构域，每个结构域约70个氨基酸，并以特定排列的6个半胱氨酸为特征。这些结构域之一也存在于马来布鲁线虫、烟草天蛾和茧蜂几丁质酶的非催化区域。围食蛋白-44在幼虫围食膜中分布均匀。逆转录聚合酶链反应检测到围食蛋白-44在所有三个幼虫龄期均有表达，但在成年绿蝇中仅检测到微量表达。该蛋白质在体外能特异性结合三-N-乙酰壳三糖和再乙酰化壳聚糖。得出的结论是，围食蛋白-44中多个富含半胱氨酸的结构域负责与围食膜的主要成分几丁质结合。围食蛋白-44可能在维持围食膜结构和确定围食膜孔隙率方面发挥作用。本报告首次对昆虫围食膜蛋白进行了表征。
