pH-induced conformational transitions of the propeptide of human cathepsin L. A role for a molten globule state in zymogen activation.
Synthesis of proteases as inactive zymogens is a very important mechanism for the regulation of their activity. For lysosomal proteases proteolytic cleavage of the propeptide is triggered by the acidic pH. By using fluorescence, circular dichroism, and NMR spectroscopy, we show that upon decreasing the pH from 6.5 to 3 the propeptide of cathepsin L loses most of the tertiary structure, but almost none of the secondary structure is lost. Another partially structured intermediate, prone to aggregation, was identified between pH 6.5 and 4. The conformation, populated below pH 4, where the activation of cathepsin L occurs, is not completely unfolded and has the properties of molten globule, including characteristic binding of the 1-anilinonaphthalene-8-sulfonic acid. This pH unfolding of the propeptide parallels a decrease of its affinity for cathepsin L and suggests the mechanism for the acidic zymogen activation. Addition of anionic polysaccharides that activate cathepsin L already at pH 5.5 unfolds the tertiary structure of the propeptide at this pH. Propeptide of human cathepsin L which is able to fold independently represents an evolutionary intermediate in the emergence of novel inhibitors originating from the enzyme proregions.
Jerala
R
R
Laboratory for Molecular Modeling and NMR Spectroscopy, National Institute of Chemistry, Hajdrihova 19, Slovenia. roman.jerala@k.si
Zerovnik
E
E
Kidric
J
J
Turk
V
V
D056668:Cathepsin L; D002403:Cathepsins / Q000737:chemistry / Q000648:ultrastructure*; D002942:Circular Dichroism; D003546:Cysteine Endopeptidases; D016264:Dextran Sulfate / Q000737:chemistry; D010450:Endopeptidases; D004789:Enzyme Activation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D011489:Protein Denaturation; D011498:Protein Precursors / Q000737:chemistry / Q000648:ultrastructure*; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins; D013050:Spectrometry, Fluorescence
D011498:Protein Precursors; D011994:Recombinant Proteins; D016264:Dextran Sulfate; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L