Fhit, a putative tumor suppressor in humans, is a dinucleoside 5',5"'-P1,P3-triphosphate hydrolase.
Human Fhit (fragile histidine triad) protein, encoded by the FHIT putative tumor suppressor gene, is a typical dinucleoside 5',5"'-P1,P3-triphosphate (Ap3A) hydrolase (EC 3.6.1.29) on the basis of its enzymatic properties we report here. Ap3A is the preferred substrate among ApnA (n = 3-6), and AMP is always one of the reaction products. Mn2+ and Mg2+ are equally stimulatory, while Zn2+ is inhibitory with Ap3A as the substrate. Values of the K(m) for Ap3A and Ap4A are 1.3 and 4.6 microM, respectively. Values of the specificity constant, kcat/K(m), for Ap3A and Ap4A are 2.0 x 10(6) and 6.7 x 10(3) s-1 M-1, respectively, for a glutathione S-transferase (GST)-Fhit fusion protein. Site-directed mutagenesis of FHIT demonstrated that all four conserved histidines are required for full activity, and the central histidine of the triad is absolutely essential for Ap3A hydrolase activity. This putative tumor suppressor is the first evidence for a connection between dinucleotide oligophosphate metabolism and tumorigenesis. Also, Fhit is the first HIT protein in which the histidine residues have been demonstrated by mutagenesis to be critical for function.
Barnes
L D
LD
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78284-7760, USA. barnesl@uthscsa.edu
Garrison
P N
PN
Siprashvili
Z
Z
Guranowski
A
A
Robinson
A K
AK
Ingram
S W
SW
Croce
C M
CM
Ohta
M
M
Huebner
K
K
D017766:Acid Anhydride Hydrolases; D001483:Base Sequence; D017931:DNA Primers; D015226:Dinucleoside Phosphates / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D016147:Genes, Tumor Suppressor; D006639:Histidine / Q000235:genetics; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013379:Substrate Specificity; D000099170:Bis(5'-adenosyl)triphosphatase
D017766:Acid Anhydride Hydrolases; D017931:DNA Primers; D015226:Dinucleoside Phosphates; D006639:Histidine; D009363:Neoplasm Proteins; D010727:Phosphoric Diester Hydrolases; D011506:Proteins; D011993:Recombinant Fusion Proteins; D000099170:Bis(5'-adenosyl)triphosphatase; C011440:diadenosine triphosphate