Effects of N‐Glycosylation Deletions on Cypridina Luciferase Activity
Effects of N‐Glycosylation Deletions on Cypridina Luciferase Activity
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DOI：
10.1111/php.12847
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发表时间：
2018-03
影响因子：
3.3
通讯作者：
R. Yasuno;Y. Mitani;Y. Ohmiya
中科院分区：
文献类型：
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作者：
R. Yasuno;Y. Mitani;Y. Ohmiya
塞浦路斯荧光素酶(Cluc)是一种从夜光藻中鉴定出来的分泌型发光蛋白，它有两个N-糖基化位点。在这项研究中，我们通过在N-糖基化的共同序列(ASN-X-Ser/Thr)上创建定点突变修饰来评估N-糖基化对Cluc性质的影响。共鉴定了8个突变体，每个由4个单残基和双残基突变体组成。突变株Cluc的产生性和相对比活力明显降低，但热稳定性和分泌效率未受影响。这些结果表明，需要N-糖基化修饰和正确的N-糖链结合位点的氨基酸序列，才能使完整的蛋白质折叠形成稳定的催化中心，或者需要底物-蛋白质相互作用残基的正确构象，或者两者都需要，糖基化修饰的缺陷与蛋白质的分泌过程和稳定性无关。
Cypridina luciferase (Cluc), a secreted luminescent protein identified from Cypridina noctiluca, has two N‐glycosylation sites. In this study, we evaluated the effects of N‐glycosylation on Cluc properties by creating site‐directed mutagenic modifications at the consensus sequence for N‐glycosylation (Asn‐X‐Ser/Thr). Eight variants consisting of four single‐ and double‐residue mutants each were characterized. The producibility and relative specific activity were apparently reduced in mutant Cluc although the thermostability and secretion efficiency were not affected. These results suggested that N‐glycosylation modifications and the proper amino acid sequence of the N‐glycan binding sites of Cluc are required for the complete protein folding to form a stable catalytic center, for the proper conformation of substrate–protein interaction residues, or for both and that defects in the glycosylation modification are not related to secretion process and stability of the protein.