中华微生物学和免疫学杂志
中華微生物學和免疫學雜誌
중화미생물학화면역학잡지
CHINESE JOURNAL OF MICROBIOLOGY AND IMMUNOLOGY
2008年
7期
616-620
,共5页
陶爱林%刘林川%王永飞%邹泽红%马三梅%赖荷%于陆%伍秋容
陶愛林%劉林川%王永飛%鄒澤紅%馬三梅%賴荷%于陸%伍鞦容
도애림%류림천%왕영비%추택홍%마삼매%뢰하%우륙%오추용
刺苋%泛过敏原%三级结构%抗原性%氨基酸置换
刺莧%汎過敏原%三級結構%抗原性%氨基痠置換
자현%범과민원%삼급결구%항원성%안기산치환
Amaranthus spinosus%Panallergen%Three dimensional structure%Antigenicity%Aminoacid replacement
目的 克隆刺苋花粉中Profilin蛋白基因,分析同源序列中不同性质氨基酸对抗原性及j级结构的贡献.方法 基于生物信息学分析已知泛过敏原Profilin的氨基酸序列并获得核心代表序列,以之为基础设计合成引物,采用Touchdown PCR技术从刺苋花粉的cDNA池中进行基因克隆,并经菌落PCR和双酶切验证;采用生物信息学软件MULTIPRED及SWISS-MODEL在线软件对所得基因编码蛋白进行抗原性评估及三级结构模拟.结果 从刺苋中获得两个序列不同的泛过敏原基因,分别命名为PRF7和PRF23.蛋白质三级结构显示:PRF7与已知的泛过敏原Profilin存在少数氨基酸的筹异,其空间结构与抗原性没有明显变化;而PRF23与北方豚草花粉泛过敏原Q64LH0之间相似程度较低,而空间结构也呈现明显的筹异;尽管Q64LH0与PRF23的伞序列抗原性均值差异无统计学意义,受一些区段上不同性质的氨基酸改变的影响,PRF23在这些区段上的抗原性显著低于Q64L-HO的抗原性.结论 基于Q64LHO和PRF23同源氨基酸序列的抗原性评估及三级结构分析,获得了不同性质氨基酸对抗原性及三级结构的贡献等信息,映射了南方花粉过敏症发牛率较低的原因所在,也为过敏原遗传改良过程中进行氨基酸置换指明了方向.
目的 剋隆刺莧花粉中Profilin蛋白基因,分析同源序列中不同性質氨基痠對抗原性及j級結構的貢獻.方法 基于生物信息學分析已知汎過敏原Profilin的氨基痠序列併穫得覈心代錶序列,以之為基礎設計閤成引物,採用Touchdown PCR技術從刺莧花粉的cDNA池中進行基因剋隆,併經菌落PCR和雙酶切驗證;採用生物信息學軟件MULTIPRED及SWISS-MODEL在線軟件對所得基因編碼蛋白進行抗原性評估及三級結構模擬.結果 從刺莧中穫得兩箇序列不同的汎過敏原基因,分彆命名為PRF7和PRF23.蛋白質三級結構顯示:PRF7與已知的汎過敏原Profilin存在少數氨基痠的籌異,其空間結構與抗原性沒有明顯變化;而PRF23與北方豚草花粉汎過敏原Q64LH0之間相似程度較低,而空間結構也呈現明顯的籌異;儘管Q64LH0與PRF23的傘序列抗原性均值差異無統計學意義,受一些區段上不同性質的氨基痠改變的影響,PRF23在這些區段上的抗原性顯著低于Q64L-HO的抗原性.結論 基于Q64LHO和PRF23同源氨基痠序列的抗原性評估及三級結構分析,穫得瞭不同性質氨基痠對抗原性及三級結構的貢獻等信息,映射瞭南方花粉過敏癥髮牛率較低的原因所在,也為過敏原遺傳改良過程中進行氨基痠置換指明瞭方嚮.
목적 극륭자현화분중Profilin단백기인,분석동원서렬중불동성질안기산대항원성급j급결구적공헌.방법 기우생물신식학분석이지범과민원Profilin적안기산서렬병획득핵심대표서렬,이지위기출설계합성인물,채용Touchdown PCR기술종자현화분적cDNA지중진행기인극륭,병경균락PCR화쌍매절험증;채용생물신식학연건MULTIPRED급SWISS-MODEL재선연건대소득기인편마단백진행항원성평고급삼급결구모의.결과 종자현중획득량개서렬불동적범과민원기인,분별명명위PRF7화PRF23.단백질삼급결구현시:PRF7여이지적범과민원Profilin존재소수안기산적주이,기공간결구여항원성몰유명현변화;이PRF23여북방돈초화분범과민원Q64LH0지간상사정도교저,이공간결구야정현명현적주이;진관Q64LH0여PRF23적산서렬항원성균치차이무통계학의의,수일사구단상불동성질적안기산개변적영향,PRF23재저사구단상적항원성현저저우Q64L-HO적항원성.결론 기우Q64LHO화PRF23동원안기산서렬적항원성평고급삼급결구분석,획득료불동성질안기산대항원성급삼급결구적공헌등신식,영사료남방화분과민증발우솔교저적원인소재,야위과민원유전개량과정중진행안기산치환지명료방향.
Objective To clone and characterize Profilin encoding genes in Amaranthus spinosus and to analyze the contribution of different amino acids in isoallergens to allergen antigenicity and tertiary structure. Methods The primers were designed according to the core sequences which were obtained by bioinformatic analysis of the known Profilin amino acid sequences, followed by gene cloning from the Ama- ranthus spinosus cDNA pool and subsequent confirmation by double-digestion, colony PCR and DNA sequen- cing. Antigenicity evaluation and tertiary structural modeling of the encoded protein were accomplished by online software MULTIPRED and SWISS-MODEL, respectively. Results Two panallergenic genes, named as PRF7 and PRF23, were acquired from Amaranthus spinosus. Sequence and structure analysis demonstra- ted that there was some discrepancy in tertiary structures of the encoded proteins, besides distinct difference in their amino acid sequences. PRF7 exhibited high homology with panallergen Profilins Q64LH0, with the identities 98%, whereas the homology of PRF23 and Q9XF42 (apple allergen) was 81%. Q64LH0 and PRF23 were modeled as 3nulA (Q42449) and lg5uB (Q9LE18), respectively. PRF23 exhibited distinct0 three dimensional structural difference in certain fragments compared with Q64LH0 and other Profilins. Though the average values of antigenicity displayed no difference between Q64LH0 and PRF23 on whole se- quences, the antigenicity of PRF23 on certain fragments was obviously lower than that of Q64LHO because of the alteration of some amino acids with different characters, implying the cause of lower incidence of hay fe- ver in South China than in North China. Conclusion Based on sequence analysis, antigenicity evaluation and tertiary structural modeling for Q64LH0 and PRF23, we obtained lots of useful information about the contribution of different amino acids to antigenicity and protein structures, thus would facilitate allergen ge- netic improvement by amino acid replacement.
