Purification and characterization of human recombinant precursor interleukin 1 beta.
作者： D Hazuda , R L Webb , P Simon , P Young
DOI: 10.1016/S0021-9258(18)94241-9
关键词:
摘要: Abstract We have purified the 31-kDa precursor of human interleukin 1 beta (proIL1 beta) from recombinant Escherichia coli expressing protein. The was characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, spectroscopy, Western blot, and for biological receptor binding activity. protein migrates at expected molecular weight on electrophoresis analytical filtration columns. specific activity is less than 10(2) units/mg in EL4 thymoma assay compared with 5 x 10(8) 17-kDa mature inactivity attributable to inability bind IL1 cells as shown competition studies using 125I-labeled beta. Inactivity not due degradation either bioactivity or assays. inactive converted an active form following proteolysis chymotrypsin which generates a carboxyl-terminal fragment 17 kDa that 6 orders magnitude more starting precursor. Removal first 114 amino acids proIL1 fully molecule. In contrast, removal 77 treatment trypsin only partially restores resultant 22-kDa exhibits 600-fold increase both activity, demonstrating direct correlation between ability sequences within pro-region inhibit receptor. Far-UV circular dichroism spectroscopy indicates similar secondary structure beta; proteins are nonhelical sheet proteins.
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