Active site of trypanothione reductase. A target for rational drug design.
The X-ray crystal structure of the enzyme trypanothione reductase, isolated from the trypanosomatid organism Crithidia fasciculata, has been solved by molecular replacement. The search model was the crystal structure of human glutathione reductase that shares approximately 40% sequence identity. The trypanosomal enzyme crystallizes in the tetragonal space group P4(1) with unit cell lengths of a = 128.9 A and c = 92.3 A. The asymmetric unit consists of a homodimer of approximate molecular mass 108 kDa. We present the structural detail of the active site as derived from the crystallographic model obtained at an intermediate stage of the analysis using diffraction data to 2.8 A resolution with an R-factor of 23.2%. This model has root-mean-square deviations from ideal geometry of 0.026 A for bond lengths and 4.7 degrees for bond angles. The trypanosomid enzyme assumes a similar biological function to glutathione reductase and, although similar in topology to human glutathione reductase, has an enlarged active site and a number of amino acid differences, steric and electrostatic, which allows it to process only the unique substrate trypanothione and not glutathione. This protein represents a prime target for chemotherapy of several debilitating tropical diseases caused by protozoan parasites belonging to the genera Trypanosoma and Leishmania. The structural differences between the parasite and host enzymes and their substrates thus provides a rational basis for the design of new drugs active against trypanosomes. In addition, our model explains the results of site-directed mutagenesis experiments, carried out on recombinant trypanothione reductase and glutathione reductases, designed by consideration of the crystal structure of human glutathione reductase.
Hunter
W N
WN
Department of Chemistry, University of Manchester, U.K.
Bailey
S
S
Habash
J
J
Harrop
S J
SJ
Helliwell
J R
JR
Aboagye-Kwarteng
T
T
Smith
K
K
Fairlamb
A H
AH
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D016831:Crithidia fasciculata / Q000201:enzymology; D003461:Crystallography; D015195:Drug Design; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism; D005980:Glutathione Reductase / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000648:ultrastructure*; D016415:Sequence Alignment; D013045:Species Specificity; D013095:Spermidine / Q000031:analogs & derivatives* / Q000378:metabolism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D014345:Trypanosoma / Q000201:enzymology; D014961:X-Ray Diffraction
C044809:trypanothione; D009247:NADH, NADPH Oxidoreductases; C049608:trypanothione reductase; D005980:Glutathione Reductase; D005978:Glutathione; D013095:Spermidine