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一种与单纯疱疹病毒糖蛋白L同源的马立克氏病病毒基因的鉴定与特性分析

Identification and characterization of a Marek's disease virus gene homologous to glycoprotein L of herpes simplex virus.

作者信息

Yoshida S, Lee L F, Yanagida N, Nazerian K

机构信息

USDA-Agricultural Research Service, Avian Disease and Oncology Laboratory, East Lansing Michigan 48823.

出版信息

Virology. 1994 Oct;204(1):414-9. doi: 10.1006/viro.1994.1546.

Abstract

We have identified three Marek's disease virus (MDV) open reading frames (ORFs) within the BamHI D fragment of MDV genome. The predicted polypeptides are homologous to UL1 (glycoprotein L, gL), UL2 (uracil-DNA glycosylase), and UL3 (nuclear localizing phosphoprotein) of herpes simplex virus type 1 (HSV-1). Comparison of the deduced amino acid sequences of these three ORFs with HSV-1 counterparts revealed overall identities of 18, 43, and 49%, respectively. In spite of the low overall amino acid identity with HSV-1 gL, the first open reading frame was identified as a gL homolog of HSV-1 based not only on the gene arrangement but also on a limited amino acid conservation among gL homologs of alpha-herpesviruses. To characterize the expression of the MDV gL gene, an antiserum to a hydrophilic region of the gene expressed in a bacterial expression vector was produced. Immunoprecipitation with this antiserum revealed a 25,000-Da polypeptide in MDV-infected cells. Furthermore, the 25,000-Da polypeptide migrated as a 18,000-Da polypeptide following PNGase F treatment. This result is consistent with the predicted molecular weight of MDV gL, considering the two potential N-glycosylation sites and the predicted N-terminal signal sequence. A recombinant fowlpox virus expressing the MDV gL gene was generated to characterize this glycoprotein. Unlike gL in MDV-infected cells, gL expressed by recFPV-gL was highly sensitive to Endo H, indicating that it was probably retained in the endoplasmic reticulum and was not properly processed to a mature form. Therefore, similar to HSV-1 coexpression and complex formation of MDV gL and gH may be required for proper processing and transport of gL to the cell surface.

摘要

我们在马立克氏病病毒(MDV)基因组的BamHI D片段中鉴定出了三个开放阅读框(ORF)。预测的多肽与单纯疱疹病毒1型(HSV-1)的UL1(糖蛋白L,gL)、UL2(尿嘧啶-DNA糖基化酶)和UL3(核定位磷蛋白)同源。将这三个ORF推导的氨基酸序列与HSV-1的对应序列进行比较,结果显示总体同源性分别为18%、43%和49%。尽管与HSV-1 gL的总体氨基酸同源性较低,但基于基因排列以及α-疱疹病毒gL同源物之间有限的氨基酸保守性,第一个开放阅读框被鉴定为HSV-1的gL同源物。为了表征MDV gL基因的表达,制备了针对在细菌表达载体中表达的该基因亲水区域的抗血清。用这种抗血清进行免疫沉淀,在MDV感染的细胞中发现了一条25,000道尔顿的多肽。此外,经PNGase F处理后,这条25,000道尔顿的多肽迁移为一条18,000道尔顿多肽。考虑到两个潜在的N-糖基化位点和预测的N端信号序列,这一结果与MDV gL预测的分子量一致。构建了表达MDV gL基因的重组禽痘病毒以表征这种糖蛋白。与MDV感染细胞中的gL不同,重组禽痘病毒表达的gL对内切糖苷酶H高度敏感,这表明它可能滞留在内质网中,没有正确加工成成熟形式。因此,与HSV-1类似,MDV gL和gH的共表达及复合物形成可能是gL正确加工并转运至细胞表面所必需的。

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