Iodoacetamide is a thiol reagent and can inhibit glycolysis and can be used to study the role of cysteine residues in enzymology. Iodoacetamide is an alkylating reagent for cysteine residues in peptide sequencing, therefore it is useful in proteomics. It is an irreversible inhibitor of enzymes with cysteine at the active site. After protein reduction, the sulfhydryls are reacted wtih an electrophile like Iodoacetamide to prevent the disulfide linkages to reform in a random manner. It can also modify methionine and histidine residues. Iodoacetamide can be used in LC-MS and LC-MS/MS protocols for proteomics research. Iodoacetamide is soluble in water, DMF and ethanol.
| Mechanism of Action | Iodoacetamide inhibits glycolysis since it irreversibly inhibit the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). This thiol reagent is used as alkylating reagent to modify thiol groups in proteins via S-carboxyamidomethylateion. |
| Molecular Formula | C2H4INO |
| Eukaryotic Cell Culture Applications | Iodoacetamide was used in astroglia-rich primary cultures of brain cells (astrocytes) and when used in concentration up to 1mM and it was found to deprive the cells of GSH, inhibit celluar GAPDH activity, lower cellular lactate production and delay cell death. |
| Electrophoresis Applications | Iodoacetamide is commonly used for alkylation of cysteine during sample prep for proteomics and subsequent polyacrylamide gel electrophoresis for protein visualization. |
| Cancer Applications | Iodoacetamide can be used in proteomics studies in cancer research. Cancer cells secrete a higher number of exosomes compared to normal cells. They carry oncogenic proteins, mRNAs, and ncRNAs which can contribute to a metastatic niche in nearby or distant cells. This study profiled the proteins in exosomes from primary tumors compared to those from metastases. Using LC-MS/MS to screen exosomal proteins, authors found significant enrichment of glycolysis enzymes mainly in the exomes from primary tumors. Studies such as these can contribute to the understanding of tumor microenvironment and cancer metastases (Kim and Cho, 2024). |
| References |
Boja ES and Fales HM (2001) Overalkylation of a protein digest with Iodoacetamide. Anal. Chem. 73 (15):3576–3582 Hains PG, Robinson PJ (2017) The impact of commonly used alkylating agents on artifactual peptide modification. J Proteome Res.16(9):3443-3447 PMID 28799334 Kim HS and Cho JY (2024) Exosome proteomes reveal glycolysis-related enzyme enrichment in primary canine mammary gland tumor compared to metastases. Proteome Sci. 22(1):4 PMID 38419074 Schmidt MM and Dringen R (2009) Differential effects of iodoacetamide and iodoacetate on glycolysis and glutathione metabolism of cultured astrocytes. Front. Neuroenerg 1:1.doi: 10.3389/neuro.14.001.2009 |