Dynamic behaviour of chromosomes during mitosis process is one of the
fundamental issues in cell biology. Although the dramatic structural changes of
chromosome during the cell cycle were observed, until last two decades more work
have got some results on proteins involved in the process and how do they function.
It’s known that the SMC proteins play crucial roles in mitotic chromosome
condensation, segregation, and DNA repair. But little work has been done on
evolution of SMCs and it is little known that what case SMCs of protists having
special evolution positon do. The thesis investigated the distribution of SMCs and
their interactional proteins in several protists and analysed the origin and evolution of
SMCs between prokaryote and eukaryote.
In this thesis, we identified whole SMCs from 4 protists using genome blast and
other bioinformatics methods. The results of analysing these seuqences showed that
all SMC homologs were found in these species except for non SMC5 homlog in
Giardia lamblia and non SMC5/SMC6 in Trypanosoma cruzi. In addtion, the results
also showed that the multiple copy of SMC homologs in Trichomonas vaginalis and
the code sequence (ORF) bias on A nuleotide. The results of coiled-coil prediction
suggested that significant difference existed between the SMCs in protists and other
eukaryotes. Especially the SMC2 and SMC4 of G. lamblia displayed unique
chracteristics of secondary structure. The characteristics may associate with the
unique chromosome dynamic changes during nuclear divission. We also investigated
the proteins interacting with SMCs and the rusults showed only three of six proteins
were found in these protists. The results suggested that the complex composed of
“SMC/Kleisin/non SMC﹠Kleisin”became simple or have divergence largely in
these protists.
In order to studying in future the function of SMC2 and TopoII, two key
factors involved in assembling the mitotic chromosome, in G. lamblia which was
found no mitotic chromosome during mitosis, the thesis has done the work that
cloning the segment of the two genes to prokaryotic expression vector and expressing in E. coli. We successfully cloned the two genes into expression vector pGEX-KG and
over expressed the fusion proteins in right E. coli. We aquired abundant inclusion
proteins and purified the inclusions. Then we used the inclusions immuned the habbit
and prepared the anti-serum.
Lastly, we studied the distribution of SMC and MukB in prokaryote and
phylogenetic relationships with eukaryote. We found MukB/MukE/MukF exsiting
only in three gama Proteobacteria, Enterobacteriales, Pasteurellaceae, Vibrionaceae
and the identity between species is very high. This results suggested the complex may
evolve recently. Non SMC or MukB were found in Chlamydiae, Rickettsiales,
epsilon Proteobacteria, and non related proteins were found. These species all
belonged to entirely parasitic bacteria. We supposed that the proteins may lose in
these species for their parasitic life and the parasites could organize their genomes
using the apparatus of hosts. The reconstructed phylogenetic trees suggested that
MukB and SMC have large distance and might undergone convergent evolution. The
SMCs of eukaryote and SMCs of Archea have more close relationships and might
share more recent common ancestor.
修改评论