Internal Structure and Preferential Protein Binding of Colloidal Aggregates

ACS Chem Biol. 2017 Jan 20;12(1):282-290. doi: 10.1021/acschembio.6b00791. Epub 2016 Dec 16.

Abstract

Colloidal aggregates of small molecules are the most common artifact in early drug discovery, sequestering and inhibiting target proteins without specificity. Understanding their structure and mechanism has been crucial to developing tools to control for, and occasionally even exploit, these particles. Unfortunately, their polydispersity and transient stability have prevented exploration of certain elementary properties, such as how they pack. Dye-stabilized colloidal aggregates exhibit enhanced homogeneity and stability when compared to conventional colloidal aggregates, enabling investigation of some of these properties. By small-angle X-ray scattering and multiangle light scattering, pair distance distribution functions suggest that the dye-stabilized colloids are filled, not hollow, spheres. Stability of the coformulated colloids enabled investigation of their preference for binding DNA, peptides, or folded proteins, and their ability to purify one from the other. The coformulated colloids showed little ability to bind DNA. Correspondingly, the colloids preferentially sequestered protein from even a 1600-fold excess of peptides that are themselves the result of a digest of the same protein. This may reflect the avidity advantage that a protein has in a surface-to-surface interaction with the colloids. For the first time, colloids could be shown to have preferences of up to 90-fold for particular proteins over others. Loaded onto the colloids, bound enzyme could be spun down, resuspended, and released back into buffer, regaining most of its activity. Implications of these observations for colloid mechanisms and utility will be considered.

MeSH terms

  • Bacterial Proteins / antagonists & inhibitors
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / metabolism
  • Colloids
  • Coloring Agents / chemistry
  • Congo Red / chemistry
  • DNA / chemistry
  • DNA / metabolism
  • Dynamic Light Scattering
  • Estradiol / analogs & derivatives
  • Estradiol / chemistry
  • Fluoresceins / chemistry
  • Fulvestrant
  • Niacinamide / analogs & derivatives
  • Niacinamide / chemistry
  • Peptides / chemistry*
  • Peptides / metabolism
  • Phenylurea Compounds / chemistry
  • Protein Binding
  • Protein Folding
  • Proteins / chemistry*
  • Proteins / metabolism
  • Ribosomal Proteins / chemistry
  • Ribosomal Proteins / metabolism
  • Scattering, Small Angle
  • Sorafenib
  • Trypsin / chemistry
  • Trypsin / metabolism
  • Trypsin Inhibitors / chemistry
  • Trypsin Inhibitors / metabolism
  • X-Ray Diffraction
  • beta-Lactamases / chemistry
  • beta-Lactamases / metabolism

Substances

  • Bacterial Proteins
  • Colloids
  • Coloring Agents
  • Fluoresceins
  • Peptides
  • Phenylurea Compounds
  • Proteins
  • Ribosomal Proteins
  • Trypsin Inhibitors
  • ribosomal protein L2
  • Fulvestrant
  • Niacinamide
  • Congo Red
  • Estradiol
  • DNA
  • Sorafenib
  • Trypsin
  • AmpC beta-lactamases
  • beta-Lactamases