title: Analyzing alkaline proteins in human colon crypt proteome.
abstract: Normal human colon crypt protein extract was resolved by two-dimensional gel electrophoresis using pH 6-11 immobilized pH gradient strips in the first dimension. The optimized isoelectric focusing protocol includes cup-loading sample application at the anode and 1.2% hydroxyethyl disulfide (DeStreak), 15% 2-propanol and 5% glycerol in the rehydration buffer. Spots were well resolved across the entire pH range up to 11. A total of 311 protein spots were identified by mass spectrometry and peptide mass mapping. After combining isoforms, 231 nonredundant proteins were grouped into 16 categories according to their subcellular locations, and 17 categories according to their physiological functions. Histone proteins, ribosomal proteins and mitochondrial proteins were among the well-resolved highest p/ proteins. Application of this protocol to the analysis of normal and neoplastic colon crypts will contribute to the proteomic study of colorectal tumorigenesis since a significant portion of the human proteins is in basic pH range.
authors.affiliation: Basic Science, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.
authors.forename: Xin-Ming
authors.initials: XM
authors.lastname: Li
authors.forename: Bhavinkumar B
authors.initials: BB
authors.lastname: Patel
authors.forename: Elena L
authors.initials: EL
authors.lastname: Blagoi
authors.forename: Maketa D
authors.initials: MD
authors.lastname: Patterson
authors.forename: Steven H
authors.initials: SH
authors.lastname: Seeholzer
authors.forename: Steven H
authors.initials: SH
authors.lastname: Seehozer
authors.forename: Tao
authors.initials: T
authors.lastname: Zhang
authors.forename: Shirish
authors.initials: S
authors.lastname: Damle
authors.forename: Zhenqiang
authors.initials: Z
authors.lastname: Gao
authors.forename: Bruce
authors.initials: B
authors.lastname: Boman
authors.forename: Anthony T
authors.initials: AT
authors.lastname: Yeung
mesh_terms: D002457:Cell Extracts / Q000737:chemistry; D003106:Colon / Q000737:chemistry* / Q000378:metabolism; D015179:Colorectal Neoplasms / Q000737:chemistry / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D011506:Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D020543:Proteome / Q000737:chemistry*
chemical_list: D002457:Cell Extracts; D011506:Proteins; D020543:Proteome