NUCLEOTIDE BINDING DOMAIN OF FRUCTOSE-1,6-BISPHOSPHATASE

被引:28
作者
CRUZ, ZM [1 ]
TANIZAKI, MM [1 ]
ELDORRY, HA [1 ]
BACILA, M [1 ]
机构
[1] UNIV SAO PAULO, INST CHEM, DEPT BIOCHEM, SAO PAULO, BRAZIL
关键词
D O I
10.1016/0003-9861(79)90516-2
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Native chicken liver fructose-1,6-bisphosphatase (Fru-P2ase) can bind to blue dextranSepharose affinity column and is not displaced by its sugar-phosphate substrate; however; it is readily eluted by the inhibitor 5′-AMP. Treatment of Fru-P2ase with pyridoxal 5′-phosphate (pyridoxal-P) in the presence of the substrate, fructose 1,6-bisphosphate, followed by reduction with NaBH4 leads to the formation of active pyridoxal-P derivatives of the enzyme showing diminished sensitivity to AMP inhibitor. The modified enzyme does not bind to the affinity column. On the other hand, in the presence of AMP modification of Fru-P2ase with pyridoxal-P occurs at the catalytic site; this modification does not alter its binding behavior toward the dye ligand. Blue dextran can also protect Fru-P2ase against AMP inhibition, and it is a competitive desensitizer for the nucleotide ligand. The results establish that blue dextran binds specifically to the allosteric site of the enzyme, and that the structure of this site may resemble that of the dinucleotide fold in other enzymes. Like native Fru-P2ase, digestion of pyridoxal-P-Fru-P2ase (with regulatory properties altered) with subtilisin causes a severalfold increase in the catalytic activity measured at pH 9.2, without significant change in the activity at pH 7.5, and produces a peptide with 56 amino acids. The residual subunit, Mr ~ 30,000, was found to contain all of the incorporated pyridoxal-P. © 1979.
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页码:424 / 433
页数:10
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