Small-Angle X-Ray Scattering of Reduced Ribonuclease A: Effects of Solution Conditions And Comparisons With a Computational Model of Unfolded Proteins
The disulfide-reduced form of bovine ribonuclease A, with the Cys thiols irreversibly blocked, was characterized by small-angle x-ray scattering. To help resolve the conflicting results and interpretations from previous studies of this model unfolded protein, we measured scattering profiles using a range of solution conditions and compared them with the profiles predicted by a computational model for a random-coil polypeptide. Analysis of the simulated and experimental profiles reveals that scattering intensities at intermediate angles, corresponding to interatomic distances in the range of 5--20 {angstrom}, are particularly sensitive to changes in solvation and can be used to assess the internal scaling behavior of the polypeptide chain, expressed as a mass fractal dimension, D{sub m}. This region of the scattering curve is also much less sensitive to experimental artifacts than is the very small angle regime (the Guinier region) that has been more typically used to characterize unfolded proteins. The experimental small-angle x-ray scattering profiles closely matched those predicted by the computational model assuming relatively small solvation energies. The scaling behavior of the polypeptide approaches that of a well-solvated polymer under conditions where it has a large net charge and at high urea concentrations. At lower urea concentrations and neutral pH, the behavior of the chain approaches that expected for {theta}-conditions, where the effects of slightly unfavorable interactions with solvent balance those of excluded volume, leading to scaling behavior comparable to that of an idealized random walk chain. Though detectable, the shift toward more compact conformations at lower urea concentrations does not correspond to a transition to a globule state and is associated with little or no reduction in conformational entropy. This type of collapse, therefore, is unlikely to greatly reduce the conformational search for the native state.
- Research Organization:
- SLAC National Accelerator Lab., Menlo Park, CA (United States)
- Sponsoring Organization:
- USDOE
- DOE Contract Number:
- AC02-76SF00515
- OSTI ID:
- 953601
- Report Number(s):
- SLAC-REPRINT-2009-282; JMOBAK; TRN: US201002%%1429
- Journal Information:
- J. Mol. Biol. 377:1576,2008, Vol. 377, Issue 5; ISSN 0022-2836
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
99 GENERAL AND MISCELLANEOUS//MATHEMATICS, COMPUTING, AND INFORMATION SCIENCE
BALANCES
BEHAVIOR
CATTLE
CHAINS
CHARGES
COMPACTS
DIAGRAMS
ENTROPY
FRACTALS
INTERACTIONS
INTERATOMIC DISTANCES
MASS
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POLYPEPTIDES
PROTEINS
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REDUCTION
RNA-ASE
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