| OBJECTIVE:The aim of the thesis is to explore the molecular function of HPS1 with the tertiary structure of it and revolutionary analysis.METHODS:The mouse HPS1 and HPS4 were cloned and deduced expression by the prokaryotic expression system.Both proteins were extracted and purified to crystallize.And the induced protein was also used to immune the rabbit to produce HPS1 antibody.Blast search for the homologous protein of HPS in the model animals.With ClustalW and phylogenetic analysis,it was supposed to predict the evolutionary status of HPS1 and its possible function.Moreover,the HPS1 and HPS2 mutant fibroblasts were constructed.With zinquin fluorescence probe,the transportation of Zn2+ could be observed.RESULTS:The murine HPS1 and HPS4 was successfully cloned.However,among those HPS1 fragments intercepted according to the hydrophilic characteristic of the amino acid,only the protein encoded by 54-753 base pairs could be deduced.Most of the deduced protein existed as the form of inclusion body and there was not enough soluble protein for crystallization.HPS4 was deduced,but the protein was also existed as the form of inclusion body.In the evolution,there were not HPS homologous protein in the lower organisms such as C.elegans,A.thaliana and yeast besides HPS2,whose homologous protein was found in lower organisms.As a result,the HPS protein was divided to two classes: one class was as AP-3,which holds homolog in yeast;the other class included proteins such as HPS1,HPS3,HPS4,HPS5,HPS6,DTNBP1,HPSS,P1dn,Cno and Mu, which exist only in the metazoan.According to the zinquin fluorescence probe,the vesicle storage of Zn2+ in ep fibroblasts was normal as that of BL6,while the amount of Zn2+ decreased in the pe fibroblasts.CONCLUSIONS:HPS is a new protein produced with the cell differentiation and the biogenesis of Lysosomal Related Organelles(LROs).AP-3 on one side and the HPS1 on the other side act at different steps in the biogenesis of LROs.
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