Molecular adaptation of Drosophila melanogaster lysozymes to a digestive function
Molecular adaptation of Drosophila melanogaster lysozymes to a digestive function
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DOI：
10.1016/s0965-1748(97)00108-2
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发表时间：
1998-05-01
影响因子：
3.8
通讯作者：
Terra, WR
中科院分区：
文献类型：
--
作者：
Regel, R;Matioli, SR;Terra, WR
用Superose柱凝胶过滤和Mono-Q柱离子交换层析，从加热的黑腹果蝇幼虫酸性提取物中纯化得到一种溶菌酶，其等电点为5.5。最终收率为67%。经十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法测定，纯化后的溶菌酶活性下降，最适pH值向酸性方向移动，Kin随培养基离子强度的升高而增大。该溶菌酶对环纹双翅目中存在的一种类似组织蛋白酶D的酶具有抗性，并显示出比鸡溶菌酶高11倍的几丁质酶活性。对纯化的溶菌酶的内肽进行显微测序表明，该酶是先前测序的Lys D基因的产物。结果表明，Lys P基因产物的等电点为7.2，最适pH在5左右，不是真正的消化酶。脊椎动物前肠发酵菌的D、黑素胃溶菌酶D和溶菌酶最显著的序列趋同是丝氨酸104和碱性氨基酸的减少，这表明这些特征对于酸性环境中的消化功能是必要的。昆虫和脊椎动物的适应性残基在酸性蛋白分解环境中具有不同的稳定性，这可能是因为它们依赖于溶菌酶的整体三维结构。最大似然系统发育和昆虫溶菌酶序列的推断表明，溶菌酶作为消化酶的招募是苍蝇的祖先条件(双翅目：环状轮虫)。(C)1998爱思唯尔科学有限公司。保留所有权利。
A lysozyme (pI 5.5) was purified to homogeneity from heated acid extracts of Drosophila melanogaster larvae, using gel filtration in a Superose column and ion-exchange chromatography in a Mono Q column. The final yield was 67%. The purified lysozyme with Mr 13 700 (determined by SDS-polyacrylamide gel electrophoresis) decreases in activity and has its pH optimum displaced towards acidic values and Kin increases as the ionic strength of the medium becomes higher. The lysozyme is resistant to a cathepsin D-like proteinase present in cyclorrhaphous Diptera and displays a chitinase activity which is 11-fold higher than that of chicken lysozyme. Microsequencing of an internal peptide of the purified lysozyme showed that this enzyme is the product of the previously sequenced Lys D gene. The results suggest that the product of the Lys P gene has pI 7.2, a pH optimum around 5 and is not a true digestive enzyme. The most remarkable sequence convergence of D, melanogaster lysozyme D and lysozymes from vertebrate foregut fermenters are serine 104 and a decrease in the number of basic amino acids, suggesting that these features are necessary for digestive function in an acid environment. Adaptive residues putatively conferring stability in an acid proteolytic environment differ between insects and vertebrates, probably because they depend on the overall three-dimensional structure of the lysozymes. A maximum likelihood phylogeny and inferences from insect lysozyme sequences showed that the recruitment of lysozymes as digestive enzymes is an ancestral condition of the flies (Diptera: Cyclorrhapha). (C) 1998 Elsevier Science Ltd. All rights reserved.