Proteome profile of cytosolic component of zebrafish liver generated by LC-ESI MS/MS combined with trypsin digestion and microwave-assisted acid hydrolysis.
The zebrafish genome has recently been sequenced and annotated allowing for high-throughput proteomic analysis. Here, we report for the first time a proteomic subset of zebrafish liver, an important organ for metabolizing toxins. Using a newly developed analytical procedure, we have identified 1204 proteins from the cytosolic component of a zebrafish liver tissue sample. Our methods involve cell-compartment fractionation of liver tissue samples, four levels of protein digestion, and off-line two-dimensional liquid chromatography (2-D LC) separations of resultant peptides. Proteins are identified using an electrospray ionization quadrupole time-of-flight tandem mass spectrometer (ESI-QTOF MS/MS), which provides high-resolution and high-accuracy mass measurement of peptide ions and their fragment ions. We demonstrate that greater proteome coverage can be achieved by combining the results obtained from four methods of protein digestion: three tryptic digests (one in buffer, one in methanol, and another in SDS), and a microwave-assisted acid hydrolysate of the protein extracts. Identified proteins--which included several groups of established protein biomarkers--were functionally classified. We discuss the functions and implications of these biomarkers within the context of zebrafish toxicology.
Wang
Nan
N
Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada.
Mackenzie
Lauren
L
De Souza
Andrea G
AG
Zhong
Hongying
H
Goss
Greg
G
Li
Liang
L
D000818:Animals; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000378:metabolism*; D006868:Hydrolysis; D008099:Liver / Q000378:metabolism*; D008872:Microwaves; D010449:Peptide Mapping / Q000379:methods; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization; D013347:Subcellular Fractions; D013997:Time Factors; D014357:Trypsin / Q000494:pharmacology*; D015027:Zebrafish
D020543:Proteome; D014357:Trypsin