- Mechanisms of catalysis and inhibition operative in the arginine deiminase from the human pathogen Giardia lamblia
-
Giardia lamblia arginine deiminase (GlAD), the topic of this paper, belongs to the hydrolase branch of the guanidine-modifying enzyme superfamily, whose members employ Cys-mediated nucleophilic catalysis to promote deimination of l-arginine and its naturally occurring derivatives. G. lamblia is the causative agent in the human disease giardiasis. The results of RNAi/antisense RNA gene-silencing studies reported herein indicate that GlAD is essential for G. lamblia trophozoite survival and thus, a potential target for the development of therapeutic agents for the treatment of giardiasis. The homodimeric recombinant protein was prepared in Escherichia coli for in-depth biochemical characterization. The 2-domain GlAD monomer consists of a N-terminal domain that shares an active site structure (depicted by an in silico model) and kinetic properties (determined by steady-state and transient state kinetic analysis) with its bacterial AD counterparts, and a C-terminal domain of unknown fold and function. GlAD was found to be active over a wide pH range and to accept l-arginine, l-arginine ethyl ester, Nα-benzoyl-l-arginine, and Nω-amino-l-arginine as substrates but not agmatine, l-homoarginine, Nα-benzoyl-l-arginine ethyl ester or a variety of arginine-containing peptides. The intermediacy of a Cys424-alkylthiouronium ion covalent enzyme adduct was demonstrated and the rate constants for formation (k1 = 80 s-1) and hydrolysis (k2 = 35 s-1) of the intermediate were determined. The comparatively lower value of the steady-state rate constant (kcat = 2.6 s-1), suggests that a step following citrulline formation is rate-limiting. Inhibition of GlAD using Cys directed agents was briefly explored. S-Nitroso-l-homocysteine was shown to be an active site directed, irreversible inhibitor whereas Nω-cyano-l-arginine did not inhibit GlAD but instead proved to be an active site directed, irreversible inhibitor of the Bacillus cereus AD.
- Li, Zhimin,Kulakova, Liudmila,Li, Ling,Galkin, Andrey,Zhao, Zhiming,Nash, Theodore E.,Mariano, Patrick S.,Herzberg, Osnat,Dunaway-Mariano, Debra
-
experimental part
p. 149 - 161
(2010/03/01)
-
- PEPTIDYL ARGININE DEIMINASE TYPE IV INHIBITOR
-
A compound represented by the general formula (I) or a salt thereof is provided: wherein R1, R2 and R3 each independently represent a hydrogen atom or an alkyl group having 1 to 3 carbon atoms, provided that at least one of R1, R2 and R3 does not represent a hydrogen atom; R4 represents an amino group which has a substituent; and R5 represents a carboxyl group which may have a substituent. Also provided is a peptidylarginine deiminase 4 inhibitor. The inhibitor can be used for the prevention and/or treatment of diseases associated with peptidylarginine deiminases (e.g., rheumatoid arthritis and multiple sclerosis).
- -
-
Page/Page column 20-21
(2008/06/13)
-