摘要
以抗人着丝粒蛋白Ｂ的单抗和多抗以及抗ＣＨＯ细胞动粒蛋白的单抗对源真核生物（ａｒｃｈｅｚｏａ）蓝氏贾第虫（Ｇｉａｒｄｉａｌａｍｂｌｉａ）和分别代表原细菌的３个枝的３种原细菌（Ｈａｌｏｂａｃｔｅｒｉｕｍ、Ｔｈｅｒｍｏｐｌａｓｍａ、Ｓｕｌｆｏｓｐｈａｅｒｅｌｌｕｓ）作了免疫电泳检查，并以小眼虫和大肠杆菌作为对照。结果表明，３种原细菌都呈阳性反应；而且贾第虫的反应情况显然比纤毛虫、眼虫、典型涡鞭毛虫、尖尾虫（Ｏｘｙｒｒｈｉｓ）等单细胞后真核生物的更接近于原细菌的情况。这不仅从一个新的方面为真核细胞起源于古代的原细菌的学说提供了新的佐证，而且从着丝粒／动粒蛋白方面证明了源真核生物贾第虫的原始性。本工作还为认识着丝粒蛋白Ｂ和动粒蛋白的起源和演化提供了线索。
To understand the structure and function of the centromere / kinetochore thoroughly,we need information on its origin and evolution, since every living organization has its ownorigin and evolutionary history. We analyzed the protein components of thecentromere / kinetochore in one archezoa, three archeabacteria and one eubacterium by western blotting with the human centromere protein B (CENP-B) monoclonal antibody,mACA-2, which recognizes the amino end region of human CENP-B; CENP-B polyclonalantibody ra-ACA-2; and mAb37A5, a monoclonal antibody against a peptide of thekinetochore in CHO cells.Giardia lamblia was used as a representative of archezoa, which possess no mitochondriaand has 70S prokaryotic ribosomes and therefore is considered the most primitive eukaryoticcell. Three archeabacteria, Halobacterium dachaidenesis sp. nov. F3, Sulksphaerellusthermoacidophilum S-5, and Thermoplasma acidophilum were used as the representatives ofeach existing group of archeabacteria-methanogenous-halophilic group, sulfur-dependentthermophilic group and thermoplasma group respectively. E. colt was used the representativeof eubacteria.On western blots using the CENP-B monoclonal antibody, mACA-2, Giardia lamblia reaCted an 80 kD band that is similar to one of the three bands tound in other protists such asEuglena gracilis and Oxyrris marina (Wu et al., 1996c). Three archeabacteria and E. coli gavenegative response to mACA-2 antibody.The CENP-B polyclonal antibody, ra-ACA-2, recognized similar 60 kD and 30 kDbands in Giardia lamblia and the three archeabacteria. The other protists tested also exhibitedthe two bands, and an additional 50 kD band (Wu et al., 1995a-c). The reference E. coltgave 60 kD band and two bands about 80 kD which were different from those of eitherarcheabacteria or giardia.Western blots of G. lamblia's extracts using the kinetochore protein monoclonalantibody, mAb37A5, Giardia lamblia gave 45 kD, 50 kD and 120 kD bands, of which the45 kD and 120 kD bands were also found in other tested protists (Wu et al., 1994). Twoarcheabacteria, Halobacterium and Sulfosphaerellus, produced 40 kD, 45 kD, and 50 kDbands. Another archeabacterium Thermoplasma gave a 35 kD band besides those three.Eubacterium E. colt did not give any positive response. From the results, we can suggest aevolutionary line of mAb37A5 antigen from archeabacteria to protists: Thermoplasma,35 kD, 40 kD, 45 kD and 50 kD; Halobacterium and Sulfospherellus, 40 kD, 45 kD and 50 kD; Giardia, 45 kD, 50 kD and 120 kD; Euglena and Oxyrris, 45 kD and 120 kD (Wu et al.,1994).Our results support the hypothesis that the eukaryotic cell evoloved from an ancient archeabacterium,and the nucleus of Giardia is indeed one of the most primitive of the eukaryotic nucleus. Therefore it is in the intermediate position betwen archeabacteria and eukaryotic cells. Our results suggest that although the centromere / kinetochore is a characteristic structure peculiar to eukaryotic cells, its protein molecular ancestors originated in the archeabacterial ancestor of dukaryotic cells. The differentiation of CENP-B's N-terminal structure most have occured after the archeabacterial stage.Acknowledegements: We would like to thank Prof. Dr. Dennis G. Searcy of Univ. Of Massachusetts, USA, for giving Thermoplasma acidophilum; Prof. Dr. William C. Earnshaw of Johns Hopkins Univ. School of Medicine, USA, for giving mACA-2 and ra-ACA-2 antibodies; Pro f. Dr. Ronald Hancock of Layal Unly., Canada, for giving mAb37A5 antibody; and Pro f. Dr. Laurence D. Etkin of the Department of Molecular Genetics,UTMDACC, USA, for the language. improvement of English manuscript.
关键词
贾第虫
原细菌
着丝粒
动粒蛋白
免疫印迹
Giardia lamblia, Archaebacteria, Origin and evloution of centromere / kineto chore proteins, Immuno-blotting
