title: Mature cathepsin L is substantially active in the ionic milieu of the extracellular medium.
abstract: The activity of cathepsin L is affected by ionic strength, resulting in the measured pH optimum being higher in acetate-4-morpholineethane sulfonic acid (MES)-Tris buffers of constant ionic strength than in phosphate buffers of constant molarity (and hence varying ionic strength). In acetate-MES-Tris and phosphate buffers of constant ionic strength across the pH range, the catalytic constant, kcat, generally peaked at ca. pH 6.5 and essentially independently of ionic strength. Km values, of ca. 5 microM, manifested a slight rising trend with increasing ionic strength, with a sharp increase to 20-25 microM, specifically at pH 6.5 and I = 0.4. At physiological ionic strengths, the specific buffer ions present affected the activity of mature cathepsin L, kcat/Km declining above pH 6.5 in phosphate buffer, but only above pH 7 in acetate-MES-Tris buffer. In Hanks' balanced salt solution, a model of the extracellular fluid, measured values at pH 7.2 were kcat, 18.9 s-1; Km, 13.5 microM; and kcat/Km, 1.4 x 10(6) M-1 s-1. The stability of cathepsin L in the physiological pH range was also differentially affected by the specific buffer ions, generally in parallel with the enzyme activity. In Hanks' balanced salt solution, mature cathepsin L was substantially active and stable, having a half-life of 179 s at pH 7.2 and 657 s at pH 6.8 (the peritumor pH).
authors.affiliation: Department of Biochemistry, University of Natal, Scottsville, Pietermaritzburg, South Africa.
authors.forename: F M
authors.initials: FM
authors.lastname: Dehrmann
authors.forename: T H
authors.initials: TH
authors.lastname: Coetzer
authors.forename: R N
authors.initials: RN
authors.lastname: Pike
authors.forename: C
authors.initials: C
authors.lastname: Dennison
mesh_terms: D000085:Acetates; D017738:Alkanesulfonic Acids; D002021:Buffers; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism*; D003546:Cysteine Endopeptidases; D010450:Endopeptidases; D004795:Enzyme Stability; D005110:Extracellular Space; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007552:Isotonic Solutions; D007668:Kidney / Q000201:enzymology; D009025:Morpholines; D009994:Osmolar Concentration; D010710:Phosphates; D014325:Tromethamine
chemical_list: D000085:Acetates; D017738:Alkanesulfonic Acids; D002021:Buffers; C072670:Hanks Balanced Salt Solution; D007552:Isotonic Solutions; D009025:Morpholines; D010710:Phosphates; D014325:Tromethamine; C004550:2-(N-morpholino)ethanesulfonic acid; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L