Interaction of Tl+ with product complexes of fructose-1,6-bisphosphatase.
Fructose-1,6-bisphosphatase requires divalent cations (Mg2+, Mn2+, or Zn2+) for catalysis, but a diverse set of monovalent cations (K+, Tl+, Rb+, or NH(4)(+)) will further enhance enzyme activity. Here, the interaction of Tl+ with fructose-1,6-bisphosphatase is explored under conditions that support catalysis. On the basis of initial velocity kinetics, Tl+ enhances catalysis by 20% with a K(a) of 1.3 mm and a Hill coefficient near unity. Crystal structures of enzyme complexes with Mg2+, Tl+, and reaction products, in which the concentration of Tl+ is 1 mm or less, reveal Mg2+ at metal sites 1, 2, and 3 of the active site, but little or no bound Tl+. Intermediate concentrations of Tl+ (5-20 mm) displace Mg2+ from site 3 and the 1-OH group of fructose 6-phosphate from in-line geometry with respect to bound orthophosphate. Loop 52-72 appears in a new conformational state, differing from its engaged conformation by disorder in residues 61-69. Tl+ does not bind to metal sites 1 or 2 in the presence of Mg2+, but does bind to four other sites with partial occupancy. Two of four Tl+ sites probably represent alternative binding sites for the site 3 catalytic Mg2+, whereas the other sites could play roles in monovalent cation activation.
Choe
Jun-Yong
JY
Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, USA.
Nelson
Scott W
SW
Fromm
Herbert J
HJ
Honzatko
Richard B
RB
D001665:Binding Sites; D002384:Catalysis; D003460:Crystallization; D006597:Fructose-Bisphosphatase / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry; D011487:Protein Conformation; D013793:Thallium / Q000494:pharmacology*
D013793:Thallium; D006597:Fructose-Bisphosphatase; D008274:Magnesium