Effect of Tween 20 on freeze-thawing- and agitation-induced aggregation of recombinant human factor XIII.

Agitation- and freeze-thawing-induced aggregation of recombinant human factor XIII (rFXIII) is due to interfacial adsorption and denaturation at the air-liquid and ice-liquid interfaces. The aggregation pathway proceeds through soluble aggregates to formation of insoluble aggregates regardless of the denaturing stimuli. A nonionic surfactant, polyoxyethylene sorbitan monolaurate (Tween 20), greatly reduces the rate of formation of insoluble aggregates as a function of surfactant concentration, thereby stabilizing native rFXIII. Maximum protection occurs at concentrations close to the critical micelle concentration (cmc), independent of initial protein concentration. To study the mechanistic aspects of the surfactant-induced stabilization, a series of spectroscopic studies were conducted. Electron paramagnetic resonance spectroscopy indicates that binding is not occurring between Tween 20 and either the native state or a folding intermediate state of rFXIII. Further, circular dichroism spectroscopy suggests that Tween 20 does not prevent the secondary structural changes induced upon guanidinium hydrochloride-induced unfolding. Taken together, these results imply that Tween 20 protects rFXIII against freeze-thawing- and agitation-induced aggregation primarily by competing with stress-induced soluble aggregates for interfaces, inhibiting subsequent transition to insoluble aggregates.

[1]  Chung C. Hsu,et al.  Protein denaturation by combined effect of shear and air-liquid interface. , 1997, Biotechnology and bioengineering.

[2]  B. Chang,et al.  Surface-induced denaturation of proteins during freezing and its inhibition by surfactants. , 1996, Journal of pharmaceutical sciences.

[3]  T. Anchordoquy,et al.  Polymers protect lactate dehydrogenase during freeze-drying by inhibiting dissociation in the frozen state. , 1996, Archives of biochemistry and biophysics.

[4]  E. Gabellieri,et al.  Proteins in frozen solutions: evidence of ice-induced partial unfolding. , 1996, Biophysical journal.

[5]  T. Arakawa,et al.  Stability of recombinant consensus interferon to air-jet and ultrasonic nebulization. , 1995, Journal of pharmaceutical sciences.

[6]  E. Blomberg,et al.  Protein interactions at solid surfaces , 1995 .

[7]  V. Vogel.  Fibronectin in a Surface-Adsorbed State: Insolubilization and Self-Assembly , 1995 .

[8]  S. Yoshioka,et al.  Stabilizing effect of amphiphilic excipients on the freeze‐thawing and freeze‐drying of lactate dehydrogenase , 1994, Biotechnology and bioengineering.

[9]  K E Avis,et al.  Freeze-thaw studies of a model protein, lactate dehydrogenase, in the presence of cryoprotectants. , 1993, Journal of parenteral science and technology : a publication of the Parenteral Drug Association.

[10]  T. Randolph,et al.  Mechanism of polyethylene glycol interaction with the molten globule folding intermediate of bovine carbonic anhydrase B. , 1992, The Journal of biological chemistry.

[11]  Charles Tanford,et al.  The Hydrophobic Effect: formation of micelles and biological membranes''John Wiley & Sons , 1991 .

[12]  N. A. Rodionova,et al.  Study of the “molten globule” intermediate state in protein folding by a hydrophobic fluorescent probe , 1991, Biopolymers.

[13]  D. Teller,et al.  Expression, purification, and characterization of human factor XIII in Saccharomyces cerevisiae. , 1990, Biochemistry.

[14]  Mitsuru Tanaka,et al.  Interaction between Hydrophilic Proteins and Nonionic Detergents Studied by Surface Tension Measurements , 1982 .

[15]  L. Hansen,et al.  Binding of the Triton X series of nonionic surfactants to bovine serum albumin. , 1980, Biochemistry.

[16]  S. Pizzo,et al.  Human Factor XIII from plasma and platelets. Molecular weights, subunit structures, proteolytic activation, and cross-linking of fibrinogen and fibrin. , 1973, The Journal of biological chemistry.

[17]  L. Stryer,et al.  The interaction of a naphthalene dye with apomyoglobin and apohemoglobin. A fluorescent probe of non-polar binding sites. , 1965, Journal of molecular biology.

[18]  L. van den Berg,et al.  Effect of freezing on the pH and composition of sodium and potassium phosphate solutions; the reciprocal system KH2PO4-Na2-HPO4-H2O. , 1959, Archives of biochemistry and biophysics.

[19]  T. Randolph,et al.  Molten Globule Intermediate of Recombinant Human Growth Hormone: Stabilization with Surfactants , 1996, Biotechnology progress.

[20]  I. Kurochkin,et al.  Domain structure, stability and domain-domain interactions in recombinant factor XIII. , 1995, Journal of molecular biology.

[21]  F. Franks,et al.  Protein destabilization at low temperatures. , 1995, Advances in protein chemistry.

[22]  P. Privalov,et al.  Cold Denaturation of Protein , 1990 .

[23]  U. Rinas,et al.  Denaturation-renaturation of the fibrin-stabilizing factor XIII a-chain isolated from human placenta. Properties of the native and reconstituted protein. , 1990, Biological chemistry Hoppe-Seyler.

[24]  F. Macritchie.  Spread monolayers of proteins. , 1986, Advances in colloid and interface science.

[25]  Joseph D. Andrade,et al.  Protein adsorption and materials biocompatibility: A tutorial review and suggested hypotheses , 1986 .

[26]  A. Helenius,et al.  Properties of detergents. , 1979, Methods in enzymology.