The structure of procaspase 6 is similar to that of active mature caspase 6.
To investigate the structural characteristics and activation mechanism of the precursor caspase, genes encoding the inactive pro-form and the active mature form of caspase 6 were expressed in Escherichia coli and the proteins of both forms were purified to homogeneity. The structure of each protein was characterized by chemical cross-linking, size-exclusion chromatography, CD and fluorescence spectroscopies. The pro-form caspase 6 exhibits a dimeric structure and its overall secondary structure was found to be similar to that of the mature caspase 6. Upon the maturation of procaspase 6, the maximum fluorescence wavelength lambda(max) was red-shifted from 330 to 337 nm and the fluorescence intensity of lambda(max) was increased. This fluorescence spectral change indicates that the environment of a tryptophan residue in the substrate-binding site can be changed to a more polar one when the procaspase 6 is processed. Taken together, our results strongly demonstrate that precursor caspase 6 exists as a dimer and its overall structure is similar to that of the active caspase 6. Our results also suggest that the local conformational change at the substrate-binding site, with no drastic change in the overall structure, seems to enable precursor caspase 6 to become the active mature enzyme.
Kang
Byoung Heon
BH
Division of Molecular and Life Sciences, National Research Laboratory of Protein Folding and Engineering, Pohang University of Science and Technology, Pohang 790-784, South Korea.
Ko
Eunsil
E
Kwon
Oh-Keun
OK
Choi
Kwan Yong
KY
D000595:Amino Acid Sequence; D001665:Binding Sites; D053178:Caspase 6; D020169:Caspases / Q000737:chemistry*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004792:Enzyme Precursors / Q000737:chemistry*; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D014364:Tryptophan
D004792:Enzyme Precursors; D011994:Recombinant Proteins; D014364:Tryptophan; D053178:Caspase 6; D020169:Caspases