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Single-molecule observation of nucleotide induced conformational changes in basal SecA-ATP hydrolysis.


ABSTRACT: SecA is the critical adenosine triphosphatase that drives preprotein transport through the translocon, SecYEG, in Escherichia coli. This process is thought to be regulated by conformational changes of specific domains of SecA, but real-time, real-space measurement of these changes is lacking. We use single-molecule atomic force microscopy (AFM) to visualize nucleotide-dependent conformations and conformational dynamics of SecA. Distinct topographical populations were observed in the presence of specific nucleotides. AFM investigations during basal adenosine triphosphate (ATP) hydrolysis revealed rapid, reversible transitions between a compact and an extended state at the ~100-ms time scale. A SecA mutant lacking the precursor-binding domain (PBD) aided interpretation. Further, the biochemical activity of SecA prepared for AFM was confirmed by tracking inorganic phosphate release. We conclude that ATP-driven dynamics are largely due to PBD motion but that other segments of SecA contribute to this motion during the transition state of the ATP hydrolysis cycle.

SUBMITTER: Chada N 

PROVIDER: S-EPMC6200364 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Single-molecule observation of nucleotide induced conformational changes in basal SecA-ATP hydrolysis.

Chada Nagaraju N   Chattrakun Kanokporn K   Marsh Brendan P BP   Mao Chunfeng C   Bariya Priya P   King Gavin M GM  

Science advances 20181024 10


SecA is the critical adenosine triphosphatase that drives preprotein transport through the translocon, SecYEG, in <i>Escherichia coli</i>. This process is thought to be regulated by conformational changes of specific domains of SecA, but real-time, real-space measurement of these changes is lacking. We use single-molecule atomic force microscopy (AFM) to visualize nucleotide-dependent conformations and conformational dynamics of SecA. Distinct topographical populations were observed in the prese  ...[more]

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