World Library  
Flag as Inappropriate
Email this Article

N-Ethylmaleimide

Article Id: WHEBN0004225577
Reproduction Date:

Title: N-Ethylmaleimide  
Author: World Heritage Encyclopedia
Language: English
Subject: Reagents, NEM, Huáng bǎi, Biochemistry, Fibronectin
Collection: Biochemistry, Biochemistry Methods, Maleimides, Reagents, Reagents for Biochemistry
Publisher: World Heritage Encyclopedia
Publication
Date:
 

N-Ethylmaleimide

N-Ethylmaleimide[1]
Identifiers
Abbreviations NEM
CAS number  YesY
PubChem
ChemSpider  YesY
UNII  N
DrugBank
KEGG  YesY
ChEBI  N
ChEMBL  YesY
Jmol-3D images Image 1
Properties
Molecular formula C6H7NO2
Molar mass 125.12528
Melting point 43 to 46 °C (109 to 115 °F; 316 to 319 K)
Boiling point 210 °C (410 °F; 483 K)
Except where noted otherwise, data are given for materials in their standard state (at 25 °C (77 °F), 100 kPa)
 N   YesY/N?)

N-Ethylmaleimide (NEM) is an maleic acid. It contains the imide functional group, but more importantly it is an alkene that is reactive toward thiols and is commonly used to modify cysteine residues in proteins and peptides.[2]

Contents

  • Organic chemistry 1
  • Case studies 2
  • References 3
  • External links 4

Organic chemistry

NEM is a Michael acceptor, which means that it adds nucleophiles such as thiols. The resulting thioether features a strong C-S bond and the reaction is virtually irreversible. Reaction with thiols occur in the pH range 6.5–7.5, NEM may react with amines or undergo hydrolysis at a more alkaline pH. NEM has been widely used to probe the functional role of thiol groups in enzymology. NEM is an irreversible inhibitor of all cysteine peptidases, with alkylation occurring at the active site thiol group (see schematic).[3][4]

Mechanism of irreversible inhibition of a cysteine peptidase with NEM.

Case studies

NEM blocks vesicular transport. In lysis buffers, 20 to 25 mM of NEM is used to inhibit de-sumoylation of proteins for Western Blot analysis. NEM has also been used as an inhibitor of deubiquitinases.

N-Ethylmaleimide was used by Arthur Kornberg and colleagues to knock out DNA polymerase III in order to compare its activity to that of DNA polymerase I (pol III and I, respectively). Kornberg had been awarded the Nobel Prize for discovering pol I, then believed to be the mechanism of bacterial DNA replication, although in this experiment he showed that pol III was the actual replicative machinery.

NEM activates ouabain-insensitive Cl-dependent K efflux in low K sheep and goat red blood cells as shown first by Peter Lauf in 1980 (A chloride dependent K+ flux induced by N ethylmaleimide in genetically low K+ sheep and goat erythrocytes.P.K. Lauf and B.E. Theg. Biochem. Biophys. Res. Comm., 92:1422, 1980). This unique discovery contributed to the molecular identification of K-Cl cotransport (KCC) in human embryonic cells transfected by KCC1 isoform cDNA, some 16 years later (Gillen CM, Brill S, Payne JA, Forbush B 3rd: Molecular cloning and functional expression of the K-Cl cotransporter from rabbit, rat, and human. A new member of the cation-chloride cotransporter family.J Biol Chem. 1996 Jul 5;271(27):16237-44). Since then NEM has been widely used as a now classic diagnostic tool to uncover or manipulate the membrane presence of K-Cl cotransport in cells of many species in the animal kingdom (Regulation of K-Cl cotransport: from function to genes. N. C. Adragna, M. Di Fulvio and P.K. Lauf, J. Membrane Biology, 200:1-29, 2004). Despite repeated unsuccessful attempts to identify chemically the target thiol group (see also: K+ Cl Cotransport: Sulfhydryl, divalent cations and the mechanism of volume activation in a red cell. P.K. Lauf. Topical Review, J. Memb. Biol. 88:1 13, 1985), it is held that, at physiological pH, NEM may form adducts with thiols within protein kinases that phosphorylate KCC at specific serine and threonine residues primarily within the C-terminal domain of the transporter (Rinehart J, Maksimova YD, Tanis JE, Stone KL, Hodson CA, Zhang J, Risinger M, Pan W, Wu D, Colangelo CM, Forbush B, Joiner CH, Gulcicek EE, Gallagher PG, Lifton RP.Cell. 2009 Aug 7;138(3):525-36. doi: 10.1016/j.cell.2009.05.031). The ensuing dephosphorylation of KCC by protein phosphatases leads to activation of KCC as first proposed by Michael Jennings in 1990 (Jennings, M. L. & Al-Rohil, N. S. J. gen. Physiol. 95, 1021−1040, 1990).

References

  1. ^ N-Ethylmaleimide at Sigma-Aldrich
  2. ^ Thiol reactive probes at Invitrogen
  3. ^ Nelson, D. L.; Cox, M. M. "Lehninger, Principles of Biochemistry" 3rd Ed. Worth Publishing: New York, 2000. ISBN 1-57259-153-6.
  4. ^ Gregory, J. D. (1955) J. Am. Chem. Soc. 77, 3922-3923

External links

  • The MEROPS online database for peptidases and their inhibitors: NEM
  • The bifunctional analogues such as p-NN'-phenylenebismaleimide can be used as cross-linking reagent for cystine residues. see Lutter, L. C., Zeichhardt, H., Kurland, C. G. & Stoffier,G. (1972) Mol. Gen. Genet. 119, 357-366.
This article was sourced from Creative Commons Attribution-ShareAlike License; additional terms may apply. World Heritage Encyclopedia content is assembled from numerous content providers, Open Access Publishing, and in compliance with The Fair Access to Science and Technology Research Act (FASTR), Wikimedia Foundation, Inc., Public Library of Science, The Encyclopedia of Life, Open Book Publishers (OBP), PubMed, U.S. National Library of Medicine, National Center for Biotechnology Information, U.S. National Library of Medicine, National Institutes of Health (NIH), U.S. Department of Health & Human Services, and USA.gov, which sources content from all federal, state, local, tribal, and territorial government publication portals (.gov, .mil, .edu). Funding for USA.gov and content contributors is made possible from the U.S. Congress, E-Government Act of 2002.
 
Crowd sourced content that is contributed to World Heritage Encyclopedia is peer reviewed and edited by our editorial staff to ensure quality scholarly research articles.
 
By using this site, you agree to the Terms of Use and Privacy Policy. World Heritage Encyclopedia™ is a registered trademark of the World Public Library Association, a non-profit organization.
 


Copyright © World Library Foundation. All rights reserved. eBooks from Project Gutenberg are sponsored by the World Library Foundation,
a 501c(4) Member's Support Non-Profit Organization, and is NOT affiliated with any governmental agency or department.