铜绿假单胞菌噬菌体PaP2的结构蛋白及其编码基因的研究

The constituent proteins of Pseudomonas aeruginosa phage PaP2 and their responding coding genes

目的 :确定铜绿假单胞菌噬菌体PaP2结构蛋白的构成及其编码基因。 方法 :CsCl梯度离心纯化噬菌体PaP2颗粒 ,SDS PAGE电泳 ,确定结构蛋白条带及其相对分子质量 ;然后电转印于PVDF膜上 ,用Edman降解法进行N端测序。根据所测的N端氨基酸残基序列在PaP2预测基因编码的蛋白质中检索 ,从而逆推其编码基因。 结果 :SDS PAGE显示PaP2含有 10条带 ,其中 1、3、4、5、8、9等 6条带的含量足以回收进行N端测序 ,测得N端氨基酸残基分别依次为 :Met Ile Glu Leu Gly、Ala Ile Ser Pro、Ala Arg Phe Ile Asp、Ser Val Tyr Ala Gly、Ala Ile Ser Arg Asn 和Ser Phe Ser Leu Gly。据此推论出它们的编码基因分别是ORF18340 2 3889、ORF4 4 19 6 4 19、ORF16 5 89 1832 2、ORF82 74 95 30、OrF15 70 8 16 6 2 5、ORF75 6 3 830 9。 结论 :噬菌体PaP2含有 10个结构蛋白 ,其中 6个主要结构蛋白的编码基因分别是orf 2 4、orf 8、orf 2 3、orf 12、orf 2 2、orf 11。

Objective:To identify the constituent proteins and their responding coding genes in Pseudomonas aeruginosa phage PaP2. Methods:Phage particles of PaP2 were purified with the method of CsCl gradient centrifugation.The phage constituent proteins separated by SDS-PAGE were transferred to a PVDF filter,six major bands were excised from the membrane and were subjected to N-terminal sequence with Edman degradation method.The coding genes of the six proteins could be identified by comparing their N-terminal sequences with amino acids sequence of the coding sequences of PaP2 genome. Results:PaP2 particles are separated to at least eight bands by SDS-PAGE. The N-terminal sequences of six proteins are in turn: Met-Ile-Glu-Leu-Gly, Ala-Ile-Ser-Pro, Ala-Arg-Phe-Ile-Asp,Ser-Val-Tyr-Ala-Gly,Ala-Ile-Ser-Arg-Asn and Ser-Phe-Ser-Leu-Gly. Their concluded coding genes are ORF18340-23889,ORF4419-6419,ORF16589-18322,ORF8274-9530,ORF15708-16625,ORF7563-8309. Conclusion:There are ten constituent proteins in PaP2 particles. Six corresponding coding genes of them are orf 24,orf 8,orf 23,orf 12,orf 22,orf 11 respectively. Their physical and chemical properties are figured out in this study.