Enzymatic Degradation of DNA Probed by in Situ X-ray Scattering

Kurinji Krishnamoorthy, Sumit Kewalramani, Ali Ehlen, Liane M. Moreau, Chad A. Mirkin*, Monica Olvera De La Cruz, Michael J. Bedzyk

*Corresponding author for this work

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

Label-free in situ X-ray scattering from protein spherical nucleic acids (Pro-SNAs, consisting of protein cores densely functionalized with covalently bound DNA) was used to elucidate the enzymatic reaction pathway for the DNase I-induced degradation of DNA. Time-course small-angle X-ray scattering (SAXS) and gel electrophoresis reveal a two-state system with time-dependent populations of intact and fully degraded DNA in the Pro-SNAs. SAXS shows that in the fully degraded state, the DNA strands forming the outer shell of the Pro-SNA were completely digested. SAXS analysis of reactions with different Pro-SNA concentrations reveals a reaction pathway characterized by a slow, rate determining DNase I-Pro-SNA association, followed by rapid DNA hydrolysis. Molecular dynamics (MD) simulations provide the distributions of monovalent and divalent ions around the Pro-SNA, relevant to the activity of DNase I. Taken together, in situ SAXS in conjunction with MD simulations yield key mechanistic and structural insights into the interaction of DNA with DNase I. The approach presented here should prove invaluable in probing other enzyme-catalyzed reactions on the nanoscale.

Original languageEnglish (US)
Pages (from-to)11382-11391
Number of pages10
JournalACS nano
Volume13
Issue number10
DOIs
StatePublished - Oct 22 2019

Keywords

  • DNA-coated proteins
  • counterion distribution
  • enzymatic DNA degradation
  • molecular dynamics simulations
  • small-angle X-ray scattering

ASJC Scopus subject areas

  • Materials Science(all)
  • Engineering(all)
  • Physics and Astronomy(all)

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