Enrichment Of Zinc Finger Proteins At The "Omic" Level

Proteomics is the study of proteins in the"omic"scale, providing comprehensive and useful information about abundance, location, chemical modification and protein–protein interactions that is complimentary to but not available from genomics. All these information can be useful for the study of disease mechanism to further explore new drug, the development of novel biomarkers for diagnosis and early detection of disease and the identification of new targets for therapeutics.Most of zinc finger proteins (ZFP) selectively bind to specific DNA sequences, play critical roles in controlling transcription of genes and involve in human carcinogenesis. The study of zinc finger proteins at the proteomic level can provide useful information for understanding the mechanism of ZFP as transcription factors, the role of ZFP on human carcinogenesis and further developing novel biomarkers for diagnosis and new drug for therapeutics of cancer.However, zinc finger proteins, as a super family of transcription factors, drop into the category of low abundant proteins, which require enrichment prior to proteomic analysis. Based on the structural properties of ZFP, three enrichment methods of ZFP are designed and discussed on this thesis, which are named as"Line","Plane"and"Space"enrichment respectively. In experiment part, it mainly focuses on confirming two assumptions of"Line"enrichment and optimizing operation conditions of enrichment method.Depending on the result of experiments, it is concluded that: acidic pH value makes the release of zinc ion easy; buffer solution affects the stability of zinc finger proteins; low concentration of DTT prevents cysteine residues from reducing and inversely high concentration chelates metal ions to destroy zinc tetrahedral structure and appropriate concentration of IA(500μM) is beneficial for minimizing the damage of ZFP and maximizing the alkylation of unprotected cysteine residues.