Primary Research Of The Relationship Between The Structure And Function Of UDP-GalNAc: Polypeptide N-Acetylgalactosaminyl Transferase 2 | Posted on:2007-10-17 | Degree:Master | Type:Thesis | Country:China | Candidate:H J Shen | Full Text:PDF | GTID:2120360185978459 | Subject:Biochemistry and Molecular Biology | Abstract/Summary: | PDF Full Text Request | Object: O-glycosylation is initiated by a family of glycosyltransferases known as the UDP-GalNAc: Polypeptide N-Acetylgalactosaminyl Transferase(EC2.4.1.41).They play an important role in tumor formation.This thesis aims to do some primary research on the relationship between the structure and function of UDP-GalNAc: Polypeptide N-Acetylgalactosaminyl Transferase 2 by yeast two hybrid systems.Methods:1.Search the protein data bank and select the template. Predict the 3D structure of ppGalNAc-T2 based on the alignment to the Templat by SWISS-MODEL and check the quality.2.Incert ppGalNAc-T2, ppGalNAc-T2F(lack ricin like domain)and ricin like domain into vector pGADT7 and verify by sequencing.Transform the vector T2-pGADT7 into the yeast strain AH109 by LiAc method and test.its activation3.Use reverse transcription polymerase chain reaction(RT-PCR) clone MMP14 hinge domain and incert it into vector pGBKT7.Test the relationship between ppGalNAc-T2 and MMP14 hinge domain by yeast two hybrid systems.Results:We predicted the 3D structure of ppGalNAc-T2. The recombination vector was constructed correctly verified by sequencing and the result of ppGalNAc-T2 reporter activation was positive.MMP14 hinge domain was cloned from SGC7901 cell and was incerted into vector pGBKT7.We verify the relationship between ppGalNAc-T2 and MMP14 hinge domain by yeast two hybrid systems,but the result was negative.Conclusion:Three distinct domain of the ppGalNAc-T2 catalytic unit was identified.The successful construction and activation test have laid a foundation for the further study in the function of ppGalNAc-T2.The...
| Keywords/Search Tags: | ppGalNAc-T, homology modeling, yeast two hybrid systems, reporter activation, MMP, gene clone, O-glycosylation | PDF Full Text Request | Related items |
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