Distinct behavior of mutant triosephosphate isomerase in hemolysate and in isolated form: molecular basis of enzyme deficiency

Abstract

In a Hungarian family with severe decrease in triosephosphate isomerase (TPI) activity, 2 germ line-identical but phenotypically differing compound heterozygote brothers inherited 2 independent (Phe240Leu and Glu145stop codon) mutations. The kinetic, thermodynamic, and associative properties of the recombinant human wild-type and Phe240Leu mutant enzymes were compared with those of TPIs in normal and deficient erythrocyte hemolysates. The specific activity of the recombinant mutant enzyme relative to the wild type was much higher (30%) than expected from the activity (3%) measured in hemolysates.

Triosephosphate isomerase deficiency: Facts and doubts

“…The functional consequence of the full-length/truncated heterodimer is not known but may result in dramatically reduced catalytic activity. This idea was supported by experimental observations, namely, isomerase activity measured from the hemolysate of a Phe240Leu/Glu145stop patient, which was 10-fold lower than the activity of human recombinant Phe240Leu mutant (30).…”

“…Table 1). Additionally, time-dependent activity decrease at elevated temperature was demonstrated for the Phe240Leu mutation (30), although it is not predicted to affect the dimer interface.…”

“…These data suggest the amino acid is not directly part of the active site but do not rule out an essential function in maintaining active site geometry (7). Dynamic modeling has revealed that the 240 mutation induced local conformational changes that do not spread to the contact surface area which could be responsible for the decrease in TPI activity (30,31). If expressed, the Glu145stop truncated protein would include the contact surface loops, as revealed by molecular dynamic modeling, resulting in a heterodimer of the full-length species and the truncated protein without energetic restriction in the compound heterozygote patient.…”

Degradation of Functional Triose Phosphate Isomerase Protein UnderliessugarkillPathology

“…Although these studies have produced important information about the disease, they have not provided much insight into neurological and degenerative aspects of TPI deficiency. Such studies have measured reduced isomerase activity from patients' erythrocytes, platelets, and lympocytes (Schneider et al 1965;Zanella et al 1985;Orosz et al 2001;Olah et al 2002Olah et al , 2005, as well as brain and muscle tissue (Ationu et al 1999). Studies have also documented markedly elevated DHAP levels, suggesting that the anemia may be caused by toxicity of DHAP or one of its catabolites (Zanella et al 1985;Eber et al 1991;Hollan et al 1997;Karg et al 2000;Olah et al 2002Olah et al , 2005.…”

“…Studies have also documented markedly elevated DHAP levels, suggesting that the anemia may be caused by toxicity of DHAP or one of its catabolites (Zanella et al 1985;Eber et al 1991;Hollan et al 1997;Karg et al 2000;Olah et al 2002Olah et al , 2005. Interestingly, DHAP levels in nonerythrocytes are only modestly increased, presumably owing to it being a substrate for phospholipid biosynthesis in other cell types, and a yeast model of TPI deficiency reports that proteins bearing the human disease mutations maintain catalytic activity (Orosz et al 2001(Orosz et al , 2006Ralser et al 2006).…”