Purification of a Peptidoglycan Recognition Protein from Hemolymph of the Silkworm, Bombyx mori*
Purification of a Peptidoglycan Recognition Protein from Hemolymph of the Silkworm, Bombyx mori*
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DOI：
10.1074/jbc.271.23.13854
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发表时间：
1996-06
期刊：
影响因子：
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通讯作者：
H. Yoshida;Kuninori Kinoshita;M. Ashida
中科院分区：
文献类型：
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作者：
H. Yoshida;Kuninori Kinoshita;M. Ashida
开发了一种用于获得均质家蚕血淋巴蛋白（肽聚糖识别蛋白，PGRP）的方法，该蛋白对肽聚糖具有亲和力并且在其与肽聚糖结合时能够触发酚氧化酶原级联反应。纯化的PGRP具有约19 kDa的分子量，并且由等电点为6.5的单一多肽组成。在不存在二价阳离子的情况下，它与肽聚糖结合，而未检测到它与β 1，3-葡聚糖和几丁质的结合。N-乙酰-D-葡糖胺-（β 1 - 4）-N-乙酰胞壁酰-L-丙氨酰-D-异谷氨酰胺不抑制纯化的PGRP与不溶性肽聚糖的结合，但片段化的可溶性肽聚糖却抑制PGRP与不溶性肽聚糖的结合。PGRP似乎需要肽聚糖作为可能的配体，以保持其聚糖部分由至少两个或更多个重复单元组成。PGRP不具有任何可检测的溶菌酶活性，其氨基酸组成和20个氨基酸残基的氨基端序列与家蚕溶菌酶不同。PGRP似乎是一种迄今未知的蛋白质。在没有PGRP的情况下，血淋巴的血浆部分中的酚氧化酶原级联不能被肽聚糖触发，这表明在它们结合后产生了某种类型的能够激活级联的活性。然而，这一活动的确切性质尚不清楚。结合肽聚糖的纯化PGRP不显著水解26种市售肽基-7-氨基-4-甲基香豆素中的任何一种，所述肽基-7-氨基-4-甲基香豆素是各种蛋白酶的底物。
A method was developed for obtaining a homogeneous silkworm hemolymph protein (peptidoglycan recognition protein, PGRP) which has affinity for peptidoglycan and the ability to trigger the prophenoloxidase cascade upon its binding to peptidoglycan. The purified PGRP had a molecular mass of about 19 kDa and is composed of a single polypeptide with an isoelectric point of 6.5. It bound to peptidoglycan in the absence of divalent cation, whereas its binding to β1,3-glucan and chitin was not detected. N-Acetyl-D-glucosaminyl-(β1-4)-N-acetylmuramyl-L-alanyl-D-isoglutamine did not inhibit purified PGRP to bind insoluble peptidoglycan, but fragmented soluble peptidoglycan did. PGRP seemed to require peptidoglycan as a possible ligand to keep its glycan portion consisting of at least two or more of the repeating unit. PGRP did not have any detectable lysozyme activity, and its amino acid composition and amino-terminal sequence of 20 amino acid residues were shown to be different from those of silkworm lysozyme. PGRP seems to be a hitherto unknown protein. In the absence of PGRP, the prophenoloxidase cascade in the plasma fraction of hemolymph could not be triggered by peptidoglycan, indicating that some type of activity, capable of activating the cascade, is generated upon their binding. However, the exact nature of this activity is not yet known. The purified PGRP bound to peptidoglycan did not hydrolyze significantly any of the 26 commercially available peptidyl-7-amino-4-methylcoumarins, substrates for various proteases.