Alterations in protein phosphorylation and glycosylation are an essential part of cell signaling mechanisms. Such changes in protein modification patterns may be directly involved in new pathways that lead to oncogenesis. It is thus essential to monitor changes in these patterns on a global scale and to be able to identify the proteins involved that may be critical in different stages of cancer progression. In order to monitor such changes on a global scale a top-down approach will be used. This strategy will involve a 2-D liquid based fractionation of intact proteins from tissue cell lysates. These fractionated proteins will be spotted onto protein microarrays for detection of phosphorylations based upon phospho-selective fluorescent dyes and antiphosphotyrosine antibodies and selective Ser/Thr phosphatases. Glycosylation patterns will also be mapped using fluorescent lectin probes on the protein microarrays. Changes in the presence of glycosylation and structural variations of glycans will be monitored using different types of lectins that are sensitive to glycan structure. Selected liquid fractionated proteins that correspond to spots that light up on the arrays will be directed into ESI-TOF MS to determine MW as a function of pi. Shifts in the pi value and the intact molecular weight of proteins identified by peptide mapping and MS/MS as compared to database values will be used together with the microarrays to map the phospho- and glyco-protein content of tumor samples. Sites of posttranslational modifications will be identified from protein digests using peptide microarrays, capillary monolithic LC-MALDI of intact proteins with on-plate digestion, RP-HPLC/ESI-LTQ MS/MS and MALDI QIT-TOF MS/MS. It will be shown that by using the total information provided by the intact MW value and ESI-MS and MALDI-TOF MS of the digests that very high sequence coverage can be obtained for any protein. An important part of this study will be to study such changes in PTMs in different stages and grades of prostate tissue and for a sufficient number of samples to search for markers that can be correlated with the progression of the disease. This research has important implications in public health in that it may provide a methodology for searching for markers of disease. These markers can be related to the stage of the disease and so may become important for personalized treatment, an important issue in clinical medicine.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM049500-14S1
Application #
8460612
Study Section
Enabling Bioanalytical and Biophysical Technologies Study Section (EBT)
Program Officer
Edmonds, Charles G
Project Start
1994-09-12
Project End
2013-04-30
Budget Start
2010-08-01
Budget End
2013-04-30
Support Year
14
Fiscal Year
2012
Total Cost
$89,990
Indirect Cost
$30,957
Name
University of Michigan Ann Arbor
Department
Surgery
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Huang, Yifan; Zhou, Shiyue; Zhu, Jianhui et al. (2017) LC-MS/MS isomeric profiling of permethylated N-glycans derived from serum haptoglobin of hepatocellular carcinoma (HCC) and cirrhotic patients. Electrophoresis 38:2160-2167
An, Mingrui; Lohse, Ines; Tan, Zhijing et al. (2017) Quantitative Proteomic Analysis of Serum Exosomes from Patients with Locally Advanced Pancreatic Cancer Undergoing Chemoradiotherapy. J Proteome Res 16:1763-1772
Tan, Zhijing; Nie, Song; McDermott, Sean P et al. (2017) Single Amino Acid Variant Profiles of Subpopulations in the MCF-7 Breast Cancer Cell Line. J Proteome Res 16:842-851
Zhu, Jianhui; Wu, Jing; Pei, Xiucong et al. (2017) Annexin A10 is a candidate marker associated with the progression of pancreatic precursor lesions to adenocarcinoma. PLoS One 12:e0175039
Yin, Haidi; Zhu, Jianhui; Wu, Jing et al. (2016) A procedure for the analysis of site-specific and structure-specific fucosylation in alpha-1-antitrypsin. Electrophoresis 37:2624-2632
Pei, Xiucong; Zhu, Jianhui; Yang, Rui et al. (2016) CD90 and CD24 Co-Expression Is Associated with Pancreatic Intraepithelial Neoplasias. PLoS One 11:e0158021
Yang, Rui; Liu, Xinhua; Thakolwiboon, Smathorn et al. (2016) Protein Markers Associated with an ALDH Sub-Population in Colorectal Cancer. J Proteomics Bioinform 9:238-247
Zhu, Jianhui; Wu, Jing; Yin, Haidi et al. (2015) Mass Spectrometric N-Glycan Analysis of Haptoglobin from Patient Serum Samples Using a 96-Well Plate Format. J Proteome Res 14:4932-9
Yin, Haidi; Tan, Zhijing; Wu, Jing et al. (2015) Mass-Selected Site-Specific Core-Fucosylation of Serum Proteins in Hepatocellular Carcinoma. J Proteome Res 14:4876-84
Nie, Song; McDermott, Sean P; Deol, Yadwinder et al. (2015) A quantitative proteomics analysis of MCF7 breast cancer stem and progenitor cell populations. Proteomics 15:3772-83

Showing the most recent 10 out of 124 publications