恶性疟原虫组氨酸富集蛋白II基因克隆及测序

CLONING AND SEQUENCING OF THE GENES CODING FOR THE HISTIDINE RICH PROTEIN II OF PLASMODIUM FALCIPARUM *

目的:比较恶性疟原虫不同分离株组氨酸富集蛋白II( HRPII)基因全编码区核苷酸序列。方法: PCR扩增恶性疟原虫海南株(FCC1/HN)和越南株(VN)的HRPII基因全编码区, 其产物经HindIII和Bam HI双酶切,定向克隆入pUC19, 采用Sanger 双脱氧链末端终止法测序。结果:FCC1/HN 株和VN 株HRPII基因均为1 020bp, 无内含子, 两者仅10 个碱基不同。FCC1/HN 株HRPII与VN 株、IMTM22 株和Itg2 株间氨基酸同源性分别为98.8% 、92.2% 和98.7% ; 4 株虫体均有拷贝数不等的丙氨酸-组氨酸-组氨酸(AHH)和丙氨酸-组氨酸-组氨酸-丙氨酸-丙氨酸-天冬氨酸(AHHAAD) 重复序列, 具有相同的信号肽和糖基化位点。4 个分离株HRPII抗原表位相同,位于易曲性较高的5端非AHH 和AHHAAD 重复区。结论: FCC1/HN 株与VN 株的HRPII高度同源,与IMTM22 株和Itg2

AIM: To compare and analyze the homology of genes encoding histidine rich proteinII (HRPII) of different Plasmodium falciparum isolates. METHODS: Using PCR technique, the complete genes coding for HRPII of P.falciparum isolates FCC1/HN and VN isolates were amplified. PCR products were digested by HindIII/BamHI and cloned into plasmid pUC19. The recombinant plasmid HRPII/pUC19 was screened and identified by PCR and restriction analysis. The cloned HRPII genes were sequenced by Sangers method. RESULTS: HRPII genes of FCC1/HN and VN isolates were successfully amplified and cloned into pUC19. DNA sequencing showed that the coding length of HRPII gene was 1 020 bp without introns in FCC1/HN and VN isolates, however, there were ten points mutations between them. FCC1/HN isolate exhibited 98 8%, 92 2% and 98 7% homology in amino acids with isolates VN, IMTM22, and Itg2, respectively. Though the numbers of repeat sequences were different in four isolates, they had the same hydrophobic leader sequence and a single putative glycosylation site. The secondary structure analysis showed that the main antigenic determinants of four isolates were located on 5end non repeat region (amino acids 1-60). CONCLUSION: FCC1/HN isolate was highly homologous in the coding region of HRPII with VN, IMTM22, and Itg2 isolate. Four isolates exhibited similar structural characteristics and antigenic determinants in HRPII.