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Characterisation of Cisplatin Binding Sites in Human Serum Proteins Using Hyphenated Multidimensional Liquid Chromatography and ESI Tandem Mass Spectrometry
Author(s) -
Will Joanna,
Wolters Dirk A.,
Sheldrick William S.
Publication year - 2008
Publication title -
chemmedchem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.817
H-Index - 100
eISSN - 1860-7187
pISSN - 1860-7179
DOI - 10.1002/cmdc.200800151
Subject(s) - chemistry , human serum albumin , tandem mass spectrometry , bovine serum albumin , liquid chromatography–mass spectrometry , methionine , trypsin , binding site , mass spectrometry , serum albumin , cysteine , tyrosine , residue (chemistry) , chromatography , cisplatin , biochemistry , plasma protein binding , amino acid , enzyme , biology , chemotherapy , genetics
Cisplatin binding sites in human serum proteins have been characterised by using combined multidimensional liquid chromatography and ESI tandem mass spectrometry (MudPIT). Following incubation periods of 3 h for cisplatin–blood serum mixtures and subsequent trypsin digestion, MS–MS spectra were recorded for individual peptides that had been separated by SCX and RP liquid chromatography. Matching of the MS–MS spectra to theoretical sequences that were generated for human proteins in the SWISS‐PROT database led to the identification of specific binding sites in human serum albumin (HSA), serotransferrin (Trfe) and other abundant serum proteins (A2mg, A1at, Apoa1, Apoa2). The cisplatin coordination sites in HSA and Trfe were confirmed by independent MudPIT studies on cisplatin reaction mixtures with the individual proteins. A total of five specific binding sites were identified for HSA, including the cysteine residue C34, two methionine sites (M329, M548) and the tyrosine and aspartate O‐donor sites Y150 (or Y148) and D375 (or E376). Methionine‐256 was established as a cisplatin coordination site for Trfe in addition to the O‐donor sites E265, Y314, E385 and T457. Inspection of the protein structures indicates that the preferred residues belong either to peripheral α helices or to flexible loops within the protein‐binding pockets. O‐donor residues dominate as cisplatin binding sites for other abundant serum proteins.