Comparative proteomic analysis of de-N-glycosylated serum from hepatitis B carriers reveals polypeptides that correlate with disease status.
Analysis of the polypeptide profile in tissues, cells, and sera by high-resolution two-dimensional (2-D) gel electrophoresis offers promise in the identification of biomarkers that correlate with disease. However, sera contain many polypeptides bearing N-linked glycosylation that can complicate interpretation. Therefore, we tested the possibility that de-N-glycosylation of the polypeptides present in human serum would result in a simplification of serum proteome profiles. Briefly, polypeptides present in human serum were left untreated or subjected to de-N-glycosylation by incubation with PNGase F and resolved by high-resolution 2-D gel electrophoresis. De-N-glycosylation reduced the number of glycoform variants, enhanced the resolution of many polypeptides and allowed other polypeptides to become visible. As an initial test of concept, clinically relevant serum samples from individuals with or without diagnosis of hepatocellular carcinoma were compared. Several polypeptides, apparent only after de-N-glycosylation, were shown to correlate with disease. Although the results are preliminary and the identities of all the putative biomarkers not yet known, the data suggest that de-N-glycosylation offers a method to enhance the resolution of serum polypeptide profiles and has value in comparative proteomic studies.
Comunale
Mary Ann
MA
Department of Biochemistry and Molecular Pharmacology, Thomas Jefferson University, The Jefferson Center, Doylestown, PA 18901, USA.
Mattu
Taj S
TS
Lowman
Melissa A
MA
Evans
Alison A
AA
London
W Thomas
WT
Semmes
O John
OJ
Ward
Michael
M
Drake
Richard
R
Romano
Patrick R
PR
Steel
Laura F
LF
Block
Timothy M
TM
Mehta
Anand
A
D000682:Amyloid / Q000737:chemistry; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D015331:Cohort Studies; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006031:Glycosylation; D006509:Hepatitis B / Q000097:blood*; D006801:Humans; D037102:Lectins / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008113:Liver Neoplasms / Q000378:metabolism; D008297:Male; D013058:Mass Spectrometry; D010455:Peptides / Q000737:chemistry*; D011134:Polysaccharides / Q000737:chemistry; D020543:Proteome; D040901:Proteomics / Q000379:methods*
D000682:Amyloid; D037102:Lectins; D010455:Peptides; D011134:Polysaccharides; D020543:Proteome