title: Facile preparation of titanium phosphate-modified chitosan for selective capture of phosphopeptides.
abstract: A facile two-step method for preparing chitosan-based immobilized metal ion affinity chromatography was developed. First, chitosan was phosphorylated by esterification with phosphoric acid, and then titanium was chelated onto the phosphorylated chitosan. The obtained chitosan-based titanium immobilized metal ion affinity chromatography was ultrafine microparticles and had good dispersibility in acidic buffer. The selectivity and sensitivity were evaluated by phosphopeptide enrichment of mixtures of α-casein and bovine serum albumin. The enriched peptides were analyzed by mass spectrum. Enrichment protocols were optimized and the optimum-loading buffer was 80% acetonitrile with 1% trifluoroacetic acid. With α-casein concentration as low as 2 pmol, 12 phosphopeptides were detected with considerably high intensity from the digest mixtures of α-casein and bovine serum albumin with molar ratio of 1:200. The microparticles was also applied in real biological samples, 29 phosphoproteins containing 40 phosphorylated sites were identified from salt-stressed Arabidopsis thaliana leaves.
authors.affiliation: Department of Plant Nutrition, College of Resources and Environmental Sciences, China Agricultural University, Beijing, P R China.
authors.forename: Feng
authors.initials: F
authors.lastname: Shen
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Hu
authors.forename: Ping
authors.initials: P
authors.lastname: Guan
authors.forename: Xueqin
authors.initials: X
authors.lastname: Ren
mesh_terms: D000327:Adsorption; D000818:Animals; D017360:Arabidopsis / Q000737:chemistry; D002417:Cattle; D048271:Chitosan / Q000737:chemistry*; D002846:Chromatography, Affinity / Q000295:instrumentation / Q000379:methods*; D010748:Phosphopeptides / Q000737:chemistry / Q000302:isolation & purification*; D010766:Phosphorylation; D018515:Plant Leaves / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000302:isolation & purification*; D014025:Titanium / Q000737:chemistry*
chemical_list: D010748:Phosphopeptides; D010940:Plant Proteins; D012710:Serum Albumin, Bovine; D048271:Chitosan; D014025:Titanium