Scripps VIVO scripps research logo

  • Index
  • Log in
  • Home
  • People
  • Organizations
  • Research
  • Events
Search form

Identification and characterization of single chain anti-cocaine catalytic antibodies

Academic Article
uri icon
  • Overview
  • Research
  • Identity
  • Additional Document Info
  • View All
scroll to property group menus

Overview

authors

  • McKenzie, K. M.
  • Mee, J. M.
  • Rogers, C. J.
  • Hixon, M. S.
  • Kaufmann, Gunnar
  • Janda, Kim

publication date

  • January 2007

journal

  • Journal of Molecular Biology  Journal

abstract

  • Cocaine is a powerful and addictive stimulant whose abuse remains a prevalent health and societal crisis. Unfortunately, no pharmacological therapies exist and therefore alternative protein-based therapies have been examined. One such approach is immunopharmacotherapy, wherein antibodies are utilized to either bind or hydrolyze cocaine thereby blocking it from exerting its euphoric effect. Towards this end, antibodies capable of binding and hydrolyzing cocaine were identified by phage display from a biased single chain antibody library generated from the spleens of mice previously immunized with a cocaine phosphonate transition state analog hapten. Two classes of antibodies emerged based on sequence homology and mode of action. Alanine scanning mutagenesis and kinetic analysis revealed that residues H97, H99, and L96 are crucial for antibodies 3F5 and 3H9 to accelerate the hydrolysis of cocaine. Antibodies 3F1 through 3F4, which are similar to our previously identified 3A6 class of antibodies, catalyze hydrolysis through transition state stabilization by tyrosine or histidine residues H50 and L94. Mutation of either one or both tyrosine residues to histidine conferred hydrolytic activity on previously inactive antibody 3F4. Mutational analysis of residue H50 of antibody 3F3 resulted in a glutamine mutant with a rate enhancement three times greater than wild-type. A double mutant, containing glutamineH50 and lysineH52, showed a tenfold rate enhancement over wild-type. These results indicate the power of initial selection of catalytic antibodies from a biased antibody library in both rapid generation and screening of mutants for improved catalysis.

subject areas

  • Alanine
  • Amino Acid Sequence
  • Animals
  • Antibodies, Catalytic
  • Cocaine
  • Complementarity Determining Regions
  • DNA Mutational Analysis
  • Esters
  • Hydrolysis
  • Kinetics
  • Mice
  • Molecular Sequence Data
  • Mutant Proteins
  • Mutation
  • Sequence Alignment
scroll to property group menus

Research

keywords

  • catalytic antibodies
  • cocaine
  • immunopharmacotherapy
  • phage display
  • structure based design
scroll to property group menus

Identity

PubMed Central ID

  • PMC1828637

International Standard Serial Number (ISSN)

  • 0022-2836

Digital Object Identifier (DOI)

  • 10.1016/j.jmb.2006.10.031

PubMed ID

  • 17084858
scroll to property group menus

Additional Document Info

start page

  • 722

end page

  • 731

volume

  • 365

issue

  • 3

©2021 The Scripps Research Institute | Terms of Use | Powered by VIVO

  • About
  • Contact Us
  • Support