Determination of sedimentation coefficients for small peptides.
Direct fitting of sedimentation velocity data with numerical solutions of the Lamm equations has been exploited to obtain sedimentation coefficients for single solutes under conditions where solvent and solution plateaus are either not available or are transient. The calculated evolution was initialized with the first experimental scan and nonlinear regression was employed to obtain best-fit values for the sedimentation and diffusion coefficients. General properties of the Lamm equations as data analysis tools were examined. This method was applied to study a set of small peptides containing amphipathic heptad repeats with the general structure Ac-YS-(AKEAAKE)nGAR-NH2, n = 2, 3, or 4. Sedimentation velocity analysis indicated single sedimenting species with sedimentation coefficients (s(20,w) values) of 0.37, 0.45, and 0.52 S, respectively, in good agreement with sedimentation coefficients predicted by hydrodynamic theory. The described approach can be applied to synthetic boundary and conventional loading experiments, and can be extended to analyze sedimentation data for both large and small macromolecules in order to define shape, heterogeneity, and state of association.
Schuck
P
P
Biomedical Engineering and Instrumentation Program, ORS, National Institutes of Health, Bethesda, Maryland 20892, USA. pschuck@helix.nih.gov
MacPhee
C E
CE
Howlett
G J
GJ
D000595:Amino Acid Sequence; D053304:Apolipoprotein C-II; D001056:Apolipoproteins C / Q000737:chemistry; D002942:Circular Dichroism; D003198:Computer Simulation; D006801:Humans; D008962:Models, Theoretical; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013329:Structure-Activity Relationship; D014461:Ultracentrifugation
D053304:Apolipoprotein C-II; D001056:Apolipoproteins C; D010446:Peptide Fragments; D010455:Peptides; C050632:polypeptide C