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The interaction between thermodynamic stability and buried free cysteines in regulating the functional half-life of fibroblast growth factor-1.

Lee J, Blaber M
J Mol Biol 393 113-27 (2009)
Related entries: 3fgm, 3fj8, 3fj9, 3fja, 3fjc, 3fjd

Cited: 20 times
EuropePMC logo PMID: 19695265

Abstract

Protein biopharmaceuticals are an important and growing area of human therapeutics; however, the intrinsic property of proteins to adopt alternative conformations (such as during protein unfolding and aggregation) presents numerous challenges, limiting their effective application as biopharmaceuticals. Using fibroblast growth factor-1 as model system, we describe a cooperative interaction between the intrinsic property of thermostability and the reactivity of buried free-cysteine residues that can substantially modulate protein functional half-life. A mutational strategy that combines elimination of buried free cysteines and secondary mutations that enhance thermostability to achieve a substantial gain in functional half-life is described. Furthermore, the implementation of this design strategy utilizing stabilizing mutations within the core region resulted in a mutant protein that is essentially indistinguishable from wild type as regard protein surface and solvent structure, thus minimizing the immunogenic potential of the mutations. This design strategy should be generally applicable to soluble globular proteins containing buried free-cysteine residues.

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  1. Disulfide bonding in protein biophysics. Fass D. Annu Rev Biophys 41 63-79 (2012)
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  3. Protein comparability assessments and potential applicability of high throughput biophysical methods and data visualization tools to compare physical stability profiles. Alsenaidy MA, Jain NK, Kim JH, Middaugh CR, Volkin DB. Front Pharmacol 5 39 (2014)
  4. New perspectives on IL-33 and IL-1 family cytokines as innate environmental sensors. Scott IC, Rees DG, Cohen ES. Biochem Soc Trans 46 1345-1353 (2018)
  5. Delivery of growth factor-based therapeutics in vascular diseases: Challenges and strategies. Xu HL, Yu WZ, Lu CT, Li XK, Zhao YZ. Biotechnol J 12 (2017)

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  1. Endocrinization of FGF1 produces a neomorphic and potent insulin sensitizer. Suh JM, Jonker JW, Ahmadian M, Goetz R, Lackey D, Osborn O, Huang Z, Liu W, Yoshihara E, van Dijk TH, Havinga R, Fan W, Yin YQ, Yu RT, Liddle C, Atkins AR, Olefsky JM, Mohammadi M, Downes M, Evans RM. Nature 513 436-439 (2014)
  2. FGF-1 Triggers Pannexin-1 Hemichannel Opening in Spinal Astrocytes of Rodents and Promotes Inflammatory Responses in Acute Spinal Cord Slices. Garré JM, Yang G, Bukauskas FF, Bennett MV. J Neurosci 36 4785-4801 (2016)
  3. Production of full-length soluble Plasmodium falciparum RH5 protein vaccine using a Drosophila melanogaster Schneider 2 stable cell line system. Hjerrild KA, Jin J, Wright KE, Brown RE, Marshall JM, Labbé GM, Silk SE, Cherry CJ, Clemmensen SB, Jørgensen T, Illingworth JJ, Alanine DG, Milne KH, Ashfield R, de Jongh WA, Douglas AD, Higgins MK, Draper SJ. Sci Rep 6 30357 (2016)
  4. A polypeptide "building block" for the β-trefoil fold identified by "top-down symmetric deconstruction". Lee J, Blaber SI, Dubey VK, Blaber M. J Mol Biol 407 744-763 (2011)
  5. An empirical phase diagram approach to investigate conformational stability of "second-generation" functional mutants of acidic fibroblast growth factor-1. Alsenaidy MA, Wang T, Kim JH, Joshi SB, Lee J, Blaber M, Volkin DB, Middaugh CR. Protein Sci 21 418-432 (2012)
  6. Structural basis of conserved cysteine in the fibroblast growth factor family: evidence for a vestigial half-cystine. Lee J, Blaber M. J Mol Biol 393 128-139 (2009)
  7. Pharmacokinetic properties of 2nd-generation fibroblast growth factor-1 mutants for therapeutic application. Xia X, Babcock JP, Blaber SI, Harper KM, Blaber M. PLoS One 7 e48210 (2012)
  8. Impact of cysteine variants on the structure, activity, and stability of recombinant human α-galactosidase A. Qiu H, Honey DM, Kingsbury JS, Park A, Boudanova E, Wei RR, Pan CQ, Edmunds T. Protein Sci 24 1401-1411 (2015)
  9. NMR characterization of copper and lipid interactions of the C2B domain of synaptotagmin I-relevance to the non-classical secretion of the human acidic fibroblast growth factor (hFGF-1). Kathir KM, Gao L, Rajalingam D, Daily AE, Brixey S, Liu H, Davis D, Adams P, Prudovsky I, Kumar TK. Biochim Biophys Acta 1798 297-302 (2010)
  10. Cooperative hydrophobic core interactions in the β-trefoil architecture. Blaber M. Protein Sci 30 956-965 (2021)
  11. Mechanism of Thimerosal-Induced Structural Destabilization of a Recombinant Rotavirus P[4] Protein Antigen Formulated as a Multi-Dose Vaccine. Kaur K, Xiong J, Sawant N, Agarwal S, Hickey JM, Holland DA, Mukhopadhyay TK, Brady JR, Dalvie NC, Tracey MK, Love KR, Love JC, Weis DD, Joshi SB, Volkin DB. J Pharm Sci 110 1054-1066 (2021)
  12. Crystal structure of type I 3-dehydroquinate dehydratase of Aquifex aeolicus suggests closing of active site flap is not essential for enzyme action. Devi AS, Ebihara A, Kuramitsu S, Yokoyama S, Kumarevel T, Ponnuraj K. Biochem Biophys Res Commun 432 350-354 (2013)
  13. Copper binding affinity of the C2B domain of synaptotagmin-1 and its potential role in the nonclassical secretion of acidic fibroblast growth factor. Jayanthi S, Kathir KM, Rajalingam D, Furr M, Daily A, Thurman R, Rutherford L, Chandrashekar R, Adams P, Prudovsky I, Kumar TK. Biochim Biophys Acta 1844 2155-2163 (2014)
  14. Expression and Refolding of the Plant Chitinase From Drosera capensis for Applications as a Sustainable and Integrated Pest Management. Sinelnikov IG, Siedhoff NE, Chulkin AM, Zorov IN, Schwaneberg U, Davari MD, Sinitsyna OA, Shcherbakova LA, Sinitsyn AP, Rozhkova AM. Front Bioeng Biotechnol 9 728501 (2021)


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