3I2G

Cocaine Esterase with mutation G173Q, bound to DTT adduct

Summary for 3I2G

• Entry DOI: 10.2210/pdb3i2g/pdb
• Related: 1ju3 1ju4 1l7q 1l7r 3I2F 3I2H 3I2I 3I2J 3I2K
• Descriptor: Cocaine esterase, CHLORIDE ION, (4S,5S)-4,5-BIS(MERCAPTOMETHYL)-1,3-DIOXOLAN-2-OL, ... (6 entities in total)
• Functional Keywords: alpha/beta hydrolase, hydrolase
• Biological source: Rhodococcus sp. MB1 'Bresler 1999'
• Cellular location: Cytoplasm : Q9L9D7
• Total number of polymer chains: 1
• Total formula weight: 64619.40

Authors

Tesmer, J.J.G.,Nance, M.R. (deposition date: 2009-06-29, release date: 2010-06-16, Last modification date: 2024-10-30)

Primary citation

Narasimhan, D.,Nance, M.R.,Gao, D.,Ko, M.C.,Macdonald, J.,Tamburi, P.,Yoon, D.,Landry, D.M.,Woods, J.H.,Zhan, C.G.,Tesmer, J.J.,Sunahara, R.K.
Structural analysis of thermostabilizing mutations of cocaine esterase.
Protein Eng.Des.Sel., 23:537-547, 2010

PubMed Abstract: Cocaine is considered to be the most addictive of all substances of abuse and mediates its effects by inhibiting monoamine transporters, primarily the dopamine transporters. There are currently no small molecules that can be used to combat its toxic and addictive properties, in part because of the difficulty of developing compounds that inhibit cocaine binding without having intrinsic effects on dopamine transport. Most of the effective cocaine inhibitors also display addictive properties. We have recently reported the use of cocaine esterase (CocE) to accelerate the removal of systemic cocaine and to prevent cocaine-induced lethality. However, wild-type CocE is relatively unstable at physiological temperatures (tau(1/2) approximately 13 min at 37 degrees C), presenting challenges for its development as a viable therapeutic agent. We applied computational approaches to predict mutations to stabilize CocE and showed that several of these have increased stability both in vitro and in vivo, with the most efficacious mutant (T172R/G173Q) extending half-life up to 370 min. Here we present novel X-ray crystallographic data on these mutants that provide a plausible model for the observed enhanced stability. We also more extensively characterize the previously reported variants and report on a new stabilizing mutant, L169K. The improved stability of these engineered CocE enzymes will have a profound influence on the use of this protein to combat cocaine-induced toxicity and addiction in humans.

PubMed: 20436035
DOI: 10.1093/protein/gzq025

Experimental method

X-RAY DIFFRACTION (2.5 Å)