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Role of disulfide bridges in determining the biological activity of interleukin 3

Clark-Lewis, Ian and Hood, Leroy E. and Kent, Stephen B. H. (1988) Role of disulfide bridges in determining the biological activity of interleukin 3. Proceedings of the National Academy of Sciences of the United States of America, 85 (21). pp. 7897-7901. ISSN 0027-8424. http://resolver.caltech.edu/CaltechAUTHORS:CLApnas88

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Abstract

Total chemical synthesis of analog proteins was used to examine the requirement for specific disulfide bridges for the biological activity of interleukin 3 (IL-3), a growth factor that stimulates multiple lineages of hemopoietic cells. Four structural analogs of the mature, 140 amino acid murine IL-3 molecule were synthesized in which specific cysteine residues were replaced by alanines. In a quantitative IL-3 assay, based on [3H]thymidine incorporation into factor-dependent cells, the IL-3 analog with alanines substituted for all four cysteines--i.e., [Ala17,79,80,140]IL-3--had 1/500th as much activity as the molecule synthesized according to the native sequence. The two analogs [Cys17,79,Ala80,140]IL-3 and [Cys17,140,Ala79,80]IL-3 had similarly low activity, whereas the [Cys17,80,Ala79,140]IL-3 analog had 2000-fold higher activity than these three analogs, and 3-fold higher than the molecule with the native sequence. This shows that in IL-3 a single disulfide bridge, between cysteines 17 and 80, is required for biological activity that approximates physiological levels. This disulfide probably stabilizes the tertiary structure of the protein to give a conformation that is optimal for function.


Item Type:Article
Additional Information:© 1988 by the National Academy of Sciences. Contributed by Leroy E. Hood, July 11, 1988. We thank J. Ringham for performing some of the IL-3 assays and M. Martin and C. Elkins for help in preparation of the manuscript. This work was supported by research grants from the National Institutes of Health, the National Science Foundation, the Monsanto Company, and Upjohn Pharmaceuticals. I.C.-L. was supported by a C.J. Martin traveling fellowship from the National Health and Medical Research Council, Canberra, Australia. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Subject Keywords:peptide synthesis; protein engineering; cysteine; lymphokine; hemopoietic growth factor
Record Number:CaltechAUTHORS:CLApnas88
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:CLApnas88
Alternative URL:http://www.pnas.org/cgi/content/abstract/85/21/7897
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:9604
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:13 Feb 2008
Last Modified:14 Nov 2014 19:20

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