N-Ethylmaleimide

N-Ethylmaleimide[1]
IUPAC name 1-Ethylpyrrole-2,5-dione
Other names Ethylmaleimide
Identifiers
Abbreviations	NEM
CAS number	128-53-0 Y
PubChem	4362
ChemSpider	4209 Y
UNII	O3C74ACM9V N
DrugBank	DB02967
KEGG	C02441 Y
ChEBI	CHEBI:44485 N
ChEMBL	CHEMBL8211 Y
Jmol-3D images	Image 1
SMILES O=C1\C=C/C(=O)N1CC
InChI InChI=1S/C6H7NO2/c1-2-7-5(8)3-4-6(7)9/h3-4H,2H2,1H3 Y Key: HDFGOPSGAURCEO-UHFFFAOYSA-N Y InChI=1/C6H7NO2/c1-2-7-5(8)3-4-6(7)9/h3-4H,2H2,1H3 Key: HDFGOPSGAURCEO-UHFFFAOYAE
Properties
Molecular formula	C6H7NO2
Molar mass	125.12528
Melting point	43-46 °C
Boiling point	210 °C, 483 K, 410 °F
N (verify) (what is: Y/N?) Except where noted otherwise, data are given for materials in their standard state (at 25 °C, 100 kPa)
Infobox references

N-Ethylmaleimide (NEM) is an organic compound that is derived from 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]

Organic chemistry

In the jargon of 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.

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.