Matrix-assisted laser desorption/ionization-MS-based relative quantification of peptides and proteins using iodoacetamide and N-methyliodoacetamide as labeling reagents.
The use of iodoacetamide (IAA) and N-methyliodoacetamide (MIAA) as labeling agents for the relative measurements of proteins using MALDI-MS is described herein. These reagents, which alkylate the thiol groups of cysteine residues in proteins, were introduced during the alkylation step of a common protein denaturation and digestion process. This approach is simpler and cheaper than those involving isotope labeling agents. The labeling agents described herein displayed good dynamic ranges and correlation coefficients for protein quantification analyses when the proteins were treated through either in-solution or in-gel digestion. The best dynamic ranges (in the molar ratio) for proteins lysozyme, transferrin, and BSA (in-solution digestion) are 0.1-10, 0.1-8, and 0.1-8, respectively. The corresponding correlation coefficients are greater than 0.99. The IAA/MIAA labeling is a useful method for the relative quantification of peptides and digested proteins when the chromatographic isotope effect is not a major concern.
Sun
Mei-Chuan
MC
Department of Chemistry, National Dong Hwa University, Hualien, Taiwan, ROC.
Chen
Cheng-Dong
CD
Huang
Yu-Shan
YS
Wu
Zhong-Siang
ZS
Ho
Yen-Peng
YP
D000595:Amino Acid Sequence; D000818:Animals; D007202:Indicators and Reagents / Q000737:chemistry; D007460:Iodoacetamide / Q000031:analogs & derivatives* / Q000737:chemistry*; D009113:Muramidase / Q000032:analysis / Q000737:chemistry; D010446:Peptide Fragments / Q000032:analysis*; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000032:analysis / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014168:Transferrin / Q000032:analysis / Q000737:chemistry
D007202:Indicators and Reagents; C091256:N-methyliodoacetamide; D010446:Peptide Fragments; D011506:Proteins; D014168:Transferrin; D012710:Serum Albumin, Bovine; D009113:Muramidase; D007460:Iodoacetamide