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Production and Characterization of an Analog of Acidic Fibroblast Growth Factor with Enhanced Stability and Biological Activity

Arakawa, Tsutomu and Horan, Thomas P. and Narhi, Linda O. and Rees, Douglas C. and Schiffer, Susan G. and Holst, Paige L. and Prestrelski, Steven J. and Tsai, Larry B. and Fox, Gary M. (1993) Production and Characterization of an Analog of Acidic Fibroblast Growth Factor with Enhanced Stability and Biological Activity. Protein Engineering, 6 (5). pp. 541-546. ISSN 0269-2139. https://resolver.caltech.edu/CaltechAUTHORS:20150205-141530315

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Abstract

We have used recombinant DNA methods to produce two forms of bovine acidic fibroblast growth factor (aFGF), one with alanine substituted for the cysteine at position 47 and the other with the Ala47 change plus the substitution of glycine for the naturally occurring histidine at position 93. Both forms were expressed at high levels in Eschtrichia coli and purified to near homogeneity by solubilization of the inclusion bodies containing the aFGF, ion exchange chromatography, refolding of the protein and hydrophobic interaction chromatography. Circular dichroic and infrared spectra suggested that the proteins are similar in secondary and tertsiary structures and contain little or no α-helical conformations. Hydrophobic interaction chromatography showed that aFGF C47A/H93G is slightly more hydrophobic than the aFGF C47A form, suggesting that residue 93 is exposed to the solvent. Half-maximal activity in an in vitro bioassay system was reached at a 10- to 20-fold lower dose for the aFGF C47A/H93G form than for the aFGF C47A form, suggesting that alteration of this residue has an effect on the region responsible for receptor binding. Addition of 50 μg/ml heparin enhanced the in vitro activity of the aFGFs, reducing the half-maximal dose to approximately 100 pg/ml for both forms, comparable to that observed previously for basic FGF with or without heparin in this assay system.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1093/protein/6.5.541 DOIArticle
http://peds.oxfordjournals.org/content/6/5/541PublisherArticle
ORCID:
AuthorORCID
Rees, Douglas C.0000-0003-4073-1185
Contact Email Address:phoebe@caltech.edu
Additional Information:© 1993 Oxford University Press. Received on October 23, 1992; revised on March 16, 1993; accepted on March 30, 1993.
Issue or Number:5
Record Number:CaltechAUTHORS:20150205-141530315
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20150205-141530315
Official Citation:Tsutomu Arakawa, Thomas P. Horan, Linda O. Narhi, Douglas C. Rees, Susan G. Schiffer, Paige L. Holst, Steven J. Prestrelski, Larry B. Tsai, and Gary M. Fox Production and characterization of an analog of acidic fibroblast growth factor with enhanced stability and biological activity Protein Eng. (1993) 6 (5): 541-546 doi:10.1093/protein/6.5.541
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:54449
Collection:CaltechAUTHORS
Deposited By: SWORD User
Deposited On:08 Feb 2015 02:46
Last Modified:03 Oct 2019 07:57

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