Molecular characterization of multivalent bioconjugates by size-exclusion chromatography with multiangle laser light scattering.
The degree of substitution and valency of bioconjugate reaction products are often poorly judged or require multiple time- and product-consuming chemical characterization methods. These aspects become critical when analyzing and optimizing the potency of costly polyvalent bioactive conjugates. In this study, size-exclusion chromatography with multiangle laser light scattering was paired with refractive index detection and ultraviolet spectroscopy (SEC-MALS-RI-UV) to characterize the reaction efficiency, degree of substitution, and valency of the products of conjugation of either peptides or proteins to a biopolymer scaffold, i.e., hyaluronic acid (HyA). Molecular characterization was more complete compared to estimates from a protein quantification assay, and exploitation of this method led to more accurate deduction of the molecular structures of polymer bioconjugates. Information obtained using this technique can improve macromolecular engineering design principles and help to better understand multivalent macromolecular interactions in biological systems.
Department of Bioengineering, University of California at Berkeley, Berkeley, California, USA.
Jacob F
JF
Pollock
Randolph S
RS
Ashton
Nikhil A
NA
Rode
David V
DV
Schaffer
Kevin E
KE
Healy
D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D002850:Chromatography, Gel / Q000379:methods*; D053823:Hedgehog Proteins / Q000737:chemistry; D006820:Hyaluronic Acid / Q000737:chemistry; D008027:Light; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D012031:Refractometry; D012542:Scattering, Radiation; D013056:Spectrophotometry, Ultraviolet
D053823:Hedgehog Proteins; D010455:Peptides; D006820:Hyaluronic Acid