Amino-acid mutations to extend the biological half-life of a therapeutically valuable mutant of human butyrylcholinesterase

Lei Fang, Shurong Hou, Liu Xue, Fang Zheng, Chang Guo Zhan

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

Cocaine is a widely abused and addictive drug without an FDA-approved medication. Our recently designed and discovered cocaine hydrolase, particularly E12-7 engineered from human butyrylcholinesterase (BChE), has the promise of becoming a valuable cocaine abuse treatment. An ideal anti-cocaine therapeutic enzyme should have not only a high catalytic efficiency against cocaine, but also a sufficiently long biological half-life. However, recombinant human BChE and the known BChE mutants have a much shorter biological half-life compared to the native human BChE. The present study aimed to extend the biological half-life of the cocaine hydrolase without changing its high catalytic activity against cocaine. Our strategy was to design possible amino-acid mutations that can introduce cross-subunit disulfide bond(s) and, thus, change the distribution of the oligomeric forms and extend the biological half-life. Three new BChE mutants (E364-532, E377-516, and E535) were predicted to have a more stable dimer structure with the desirable cross-subunit disulfide bond(s) and, therefore, a different distribution of the oligomeric forms and a prolonged biological half-life. The rational design was followed by experimental tests in vitro and in vivo, confirming that the rationally designed new BChE mutants, i.e. E364-532, E377-516, and E535, indeed had a remarkably different distribution of the oligomeric forms and prolonged biological half-life in rats from ∼7 to ∼13 h without significantly changing the catalytic activity against (-)-cocaine. This is the first demonstration that rationally designed amino-acid mutations can significantly prolong the biological half-life of a high-activity enzyme without significantly changing the catalytic activity.

Original languageEnglish
Pages (from-to)18-25
Number of pages8
JournalChemico-Biological Interactions
Volume214
Issue number1
DOIs
StatePublished - May 5 2014

Bibliographical note

Funding Information:
This work was supported by the NIH (Grants R01 DA035552 , R01 DA032910 , R01 DA013930 , and R01 DA025100 to Zhan). The authors also acknowledge the Computer Center at the University of Kentucky for supercomputing time on a Dell X-series Cluster with 384 nodes or 4,768 processors.

Keywords

  • Cholinesterase
  • Cocaine
  • Enzyme therapy
  • Molecular modeling

ASJC Scopus subject areas

  • Toxicology

Fingerprint

Dive into the research topics of 'Amino-acid mutations to extend the biological half-life of a therapeutically valuable mutant of human butyrylcholinesterase'. Together they form a unique fingerprint.

Cite this