Thermodynamic analysis of the binding of the polyglutamate chain of 5-formyltetrahydropteroylpolyglutamates to serine hydroxymethyltransferase

Biochemistry. 1998 Sep 29;37(39):13536-42. doi: 10.1021/bi980827u.


The thermodynamic parameters for the binding of 5-formyltetrahydrofolate (5-CHO-H4PteGlun) and its polyglutamate forms to rabbit liver cytosolic serine hydroxymethyltransferase (SHMT) were determined by a combination of isothermal titration calorimetry and spectrophotometry. Binding of 5-CHO-H4PteGlun to SHMT exhibits both positive enthalpy and entropy, showing that binding is entropically driven. 5-CHO-H4PteGlu5 has a 300-fold increased affinity for SHMT compared to 5-CHO-H4PteGlu. This increase in affinity is due primarily to a decrease in the positive enthalpy with little change in entropy. A variety of anions inhibit the binding of 5-CHO-H4PteGlu5 with Ki values in the 10-20 mM range. Anions are ineffective inhibitors of 5-CHO-H4PteGlu binding to SHMT, showing that anions compete for the polyglutamate binding site. There was little difference in the Ki values for a series of dicarboxylic acids as inhibitors of 5-CHO-H4PteGlu5, suggesting that spacing of the negative charges may not be important in determining their effectiveness as inhibitors. Both the mono- and pentaglutamate derivatives of 5-CHO-H4PteGlun were cross-linked to SHMT by a carbodiimide reaction to Lys-450 which resides in a stretch of Lys, His, and Arg residues.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Calorimetry
  • Formyltetrahydrofolates / chemistry*
  • Glycine Hydroxymethyltransferase / chemistry*
  • Glycine Hydroxymethyltransferase / metabolism
  • Humans
  • Liver / enzymology
  • Molecular Sequence Data
  • Osmolar Concentration
  • Polyglutamic Acid / chemistry*
  • Polyglutamic Acid / metabolism
  • Rabbits
  • Spectrophotometry, Ultraviolet
  • Tetrahydrofolates / chemistry
  • Thermodynamics


  • Formyltetrahydrofolates
  • Tetrahydrofolates
  • Polyglutamic Acid
  • Glycine Hydroxymethyltransferase
  • 5-methyltetrahydrofolate