title: Nonnative protein polymers: structure, morphology, and relation to nucleation and growth.
abstract: Thermally induced aggregates of alpha-chymotrypsinogen A and bovine granulocyte-colony stimulating factor in acidic solutions were characterized by a combination of static and dynamic light scattering, spectroscopy, transmission electron microscopy, and monomer loss kinetics. The resulting soluble, high-molecular weight aggregates (approximately 10(3)-10(5) kDa) are linear, semiflexible polymer chains that do not appreciably associate with one another under the conditions at which they were formed, with classic power-law scaling of the radius of gyration and hydrodynamic radius with weight-average molecular weight (M(w)). Aggregates in both systems are composed of nonnative monomers with elevated levels of beta-sheet secondary structure, and bind thioflavine T. In general, the aggregate size distributions showed low polydispersity by light scattering. Together with the inverse scaling of M(w) with protein concentration, the results clearly indicate that aggregation proceeds via nucleated (chain) polymerization. For alpha-chymotrypsinogen A, the scaling behavior is combined with the kinetics of aggregation to deduce separate values for the characteristic timescales for nucleation (tau(n)) and growth (tau(g)), as well as the stoichiometry of the nucleus (x). The analysis illustrates a general procedure to noninvasively and quantitatively determine tau(n), tau(g), and x for soluble (chain polymer) aggregates, as well as the relationship between tau(n)/tau(g) and aggregate M(w).
authors: Department of Chemical Engineering, University of Delaware, Newark, Delaware, USA.
authors: William F
authors: WF
authors: Weiss
authors: Travis K
authors: TK
authors: Hodgdon
authors: Eric W
authors: EW
authors: Kaler
authors: Abraham M
authors: AM
authors: Lenhoff
authors: Christopher J
authors: CJ
authors: Roberts
mesh_terms: D002918:Chymotrypsin / Q000737:chemistry* / Q000648:ultrastructure*; D003198:Computer Simulation; D003460:Crystallization / Q000379:methods*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000737:chemistry* / Q000648:ultrastructure*; D008956:Models, Chemical; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000648:ultrastructure*; D011487:Protein Conformation
chemical_list: D046912:Multiprotein Complexes; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D002918:Chymotrypsin; C117608:alpha-chymotrypsin
references: Biotechnol Bioeng. 2007 Jan 1;96(1):1-8
references: 17136749
references: J Mol Biol. 1993 Dec 5;234(3):640-53
references: 7504736
references: Protein Sci. 1993 Mar;2(3):404-10
references: 8453378
references: J Mol Biol. 2004 Sep 3;342(1):345-53
references: 15313629
references: Fold Des. 1998;3(1):R9-23
references: 9502314
references: Anal Biochem. 2006 Jun 1;353(1):144-6
references: 16620754
references: Biotechnol Bioeng. 1997 May 20;54(4):333-43
references: 18634100
references: J Pharm Sci. 2003 May;92(5):1095-111
references: 12712430
references: Protein Sci. 2006 May;15(5):1133-41
references: 16597825
references: Microsc Res Tech. 2005 Jul;67(3-4):210-7
references: 16103997
references: Biophys J. 2001 Sep;81(3):1805-22
references: 11509390
references: Curr Opin Struct Biol. 2006 Feb;16(1):118-26
references: 16434184
references: J Protein Chem. 2002 Mar;21(3):137-43
references: 12018614
references: Acc Chem Res. 2006 Sep;39(9):671-9
references: 16981684
references: Trends Genet. 2006 Aug;22(8):408-11
references: 16806565
references: J Biol Chem. 1999 Jun 25;274(26):18589-96
references: 10373469
references: AAPS J. 2006;8(3):E501-7
references: 17025268
references: Biochemistry. 2002 Oct 15;41(41):12546-51
references: 12369846
references: Biochemistry. 2007 Jun 26;46(25):7365-73
references: 17536835
references: Blood. 1988 Dec;72(6):2074-81
references: 3264199
references: J Biol Chem. 2004 Jul 23;279(30):31374-82
references: 15133040
references: Pharm Dev Technol. 2001 Aug;6(3):441-7
references: 11485185
references: J Pharm Sci. 2003 Sep;92(9):1793-804
references: 12949998
references: Biophys Chem. 1995 Jun-Jul;55(1-2):69-93
references: 17020868
references: Biochemistry. 2007 Jun 19;46(24):7079-87
references: 17521170
references: Biochemistry. 2001 Jan 16;40(2):586-95
references: 11148054
references: Nature. 2002 Apr 4;416(6880):507-11
references: 11932737
references: Proc Natl Acad Sci U S A. 2001 Feb 27;98(5):2375-80
references: 11226247
references: Semin Cell Dev Biol. 2004 Feb;15(1):3-16
references: 15036202
references: Nature. 2005 Sep 8;437(7056):266-9
references: 16148936
references: Biophys J. 1992 Apr;61(4):921-35
references: 19431826
references: Q Rev Biophys. 1998 Feb;31(1):1-39
references: 9717197
references: Eur Biophys J. 2003 Dec;32(8):710-23
references: 12898068
references: Q Rev Biophys. 2006 Feb;39(1):1-55
references: 16772049
references: J Biol Chem. 1996 Nov 22;271(47):29525-8
references: 8939877
references: Biochim Biophys Acta. 2004 May 6;1698(2):131-53
references: 15134647
references: J Phys Chem B. 2007 Jul 12;111(27):7897-913
references: 17571872
references: J Biol Chem. 2004 Apr 23;279(17):17490-9
references: 14752113
references: J Pharm Sci. 2005 Dec;94(12):2735-48
references: 16258998
references: Nature. 1964 Mar 28;201:1284-7
references: 14151403
references: J Mol Biol. 2000 Jul 7;300(2):235-9
references: 10873462
references: Proc Natl Acad Sci U S A. 2005 Oct 25;102(43):15400-5
references: 16230628
references: J Clin Pathol. 1969 Jul;22(4):410-3
references: 5798627
references: Biochemistry. 2007 Jun 26;46(25):7558-71
references: 17530865
references: Appl Opt. 2001 Aug 20;40(24):4087-91
references: 18360445
references: Biochemistry. 2006 Dec 26;45(51):15702-9
references: 17176092
references: Protein Sci. 2004 Nov;13(11):3017-27
references: 15459333
references: Biochim Biophys Acta. 2004 Sep 6;1690(1):33-41
references: 15337168
references: Biochemistry (Mosc). 2002 May;67(5):525-33
references: 12059771
references: Protein Pept Lett. 2006;13(3):219-27
references: 16515449
references: Int J Pharm. 2005 Jan 31;289(1-2):1-30
references: 15652195
references: Biophys J. 1986 Oct;50(4):583-99
references: 3779001
references: Pharm Res. 2003 Sep;20(9):1325-36
references: 14567625
references: Biochemistry. 2005 Jun 21;44(24):8898-907
references: 15952797
references: Biochemistry. 2001 Mar 27;40(12):3525-35
references: 11297418
references: Methods Enzymol. 1999;309:256-74
references: 10507029
references: Q Rev Biophys. 2005 Aug;38(3):245-88
references: 16780604