Abstract
Interleukin-1 beta (IL-1 beta) is derived from an inactive precursor by proteolytic cleavage. To study IL-1 beta processing, we expressed the precursor in Escherichia coli, partially purified it, and used it as a substrate for various potentially relevant protease preparations. The precursor alone was virtually inactive, but incubation with membranes from human monocytes or myeloid cell lines yielded a 500-fold increase in IL-1 bioactivity. Western blot analysis of the incubated material showed that the 31,000-Da precursor is broken down to three major products, ranging from 17,400 to about 19,000 Da. The most active of these products is the smallest one, and it co-migrates during electrophoresis with mature IL-1 beta. Four purified known proteases were also tested for their effect on precursor IL-1 beta, and none of these products co-migrated with the mature protein. Chymotrypsin and Staphylococcus aureus protease yielded slightly larger products, which were highly active. Elastase and trypsin yielded substantially larger products, and these had little IL-1 activity. The products of three of the known proteases were identified by NH2-terminal sequencing. These results show conclusively that proteolysis of precursor IL-1 beta generates biological activity and that the cleavage must occur close to the mature NH2 terminus.
Highlights
Interleukin-la (IL-18) is derived from an inactive 1/3consist of approximately the COOH-terminal halves of precursorbyproteolytic cleavage
Processing, we expressed the precursor Einscherichia terminal half appears to be essential for biological activity: coli, partially purifiedit, and used itas a substrate for when IL-la and IL-10 mRNAs are translated in vitro, only variouspotentiallyrelevantproteasepreparations
The extracts were diluted 1:4 to establish conditionsunder which the r-precursor IL-10 bound to theQ-Sepharose column: a reduction of the saltconcentration to less than 50 mM and, for the urea extract, areduction of the urea concentration to 2 M
Summary
The products of three of the known proteasefosrms remain unknown Our approach to this problem has were identified byNHz-terminalsequencing.These been to express the precursor of IL-10 in Escherichia coli, resultsshow conclusively that proteolysoifs precursor partially purify it, and use it as a substrate for extracts of IL-la generates biologicalactivity and that thecleavage must occur close to the matuNreH2 terminus. Or Expression of r-Precursor IL-lfi in E. coli-Recombinant precursor indirectly, IL-1 plays a role in theactivation of hematopoietic IL-10 was expressed in E. coli under the control of the phage X PL stem cells and T-cells, in induction of fever and acute phase proteins, in degradation of cartilage, in inflammation, and in wound healing Both of the IL-1s have a molecular mass of about 17,400 Da, but cloning of the cDNAs for these proteins indicated that the initial translation products should have molecular promoter and ~1857’”thermolabile repressor [11]. Asp ser arq g/v ser met ala glu val pro glu leu ala GAC TCT AGA GGATCC TAqGTAAGGAGqTTTAACC? ATG G,CA GAAGTA CCG F A G CTC,GCC; I
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
