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A proteomic approach to identify digestive enzymes, their exocytic and microapocrine secretory routes and their compartmentalization in the midgut of Spodoptera frugiperda
Comparative Biochemistry and Physiology B: Biochemistry & Molecular Biology ( IF 1.9 ) Pub Date : 2021-08-24 , DOI: 10.1016/j.cbpb.2021.110670
Felipe J Fuzita 1 , Giuseppe Palmisano 2 , Daniel C Pimenta 3 , Walter R Terra 1 , Clélia Ferreira 1
Affiliation  

A proteomic approach was used to identify the digestive enzymes secreted by exocytosis and by microapocrine vesicles and enzyme midgut compartmentalization in Spodoptera frugiperda larvae. For this, proteomic analyses were performed in isolated midgut enterocyte microvillar membrane, in a fraction enriched in microapocrine vesicles (separated in soluble and membrane fractions), in the washings of the peritrophic membrane to isolate its loosely- and tightly-bound proteins, and in the peritrophic membrane contents. PM washings correspond to proteins extracted from the mucus layer surrounding PM. Serine endopeptidases (trypsins, chymotrypsins and serine endopeptidase homologs that have substitutions in the catalytic residues) and lipases are mainly secreted by exocytosis. Aminopeptidases are mainly microvillar enzymes and some are secreted membrane-bound to microapocrine vesicles, whereas carboxypeptidase isoforms follow different secretory routes. The results also showed that most polymer hydrolases (such as amylase and endopeptidases) are not retained in the ectoperitrophic fluid (found in PM washings but absent from PM contents). On the contrary, most enzymes involved in intermediate digestion (exemplified by carboxypeptidase and aminopeptidase) do not pass through the peritrophic membrane. Finally, the data revealed that the protein composition of PM includes peritrophins classified as peritrophic membrane proteins, PMP, and chitin deacetylase.



中文翻译:

一种识别消化酶的蛋白质组学方法,它们的胞外分泌和微量分泌途径及其在草地贪夜蛾中肠中的区室化

蛋白质组学方法用于鉴定草地贪夜蛾胞吐作用和微大分泌囊泡分泌的消化酶和酶中肠区室化幼虫。为此,在分离的中肠肠细胞微绒毛膜、富含微大分泌囊泡的部分(分离为可溶性和膜部分)、围营养膜的洗涤液中进行蛋白质组学分析以分离其松散和紧密结合的蛋白质,以及围营养膜内容物。PM 洗涤物对应于从 PM 周围的粘液层中提取的蛋白质。丝氨酸内肽酶(胰蛋白酶、糜蛋白酶和丝氨酸内肽酶同系物在催化残基中有取代)和脂肪酶主要通过胞吐作用分泌。氨基肽酶主要是微绒毛酶,有些是分泌的膜结合到微大汗腺泡,而羧肽酶同种型遵循不同的分泌途径。结果还表明,大多数聚合物水解酶(如淀粉酶和内肽酶)不保留在外营养液中(在 PM 洗涤液中发现但在 PM 内容物中不存在)。相反,大多数参与中间消化的酶(例如羧肽酶和氨肽酶)不通过围营养膜。最后,数据显示,PM 的蛋白质组成包括归类为围养膜蛋白、PMP 和几丁质脱乙酰酶的围养蛋白。

更新日期:2021-09-01
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