Dynamic Fluctuations Provide the Basis of a Conformational Switch Mechanism in Apo Cyclic AMP Receptor Protein

Burcu Aykaç Fas,Yusuf Tutar,Türkan Haliloğlu,Dennis R Livesay

doi:10.1371/journal.pcbi.1003141

Burcu Aykaç Fas, Yusuf Tutar + Show 2 more

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https://doi.org/10.1371/journal.pcbi.1003141

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Abstract

Escherichia coli cyclic AMP Receptor Protein (CRP) undergoes conformational changes with cAMP binding and allosterically promotes CRP to bind specifically to the DNA. In that, the structural and dynamic properties of apo CRP prior to cAMP binding are of interest for the comprehension of the activation mechanism. Here, the dynamics of apo CRP monomer/dimer and holo CRP dimer were studied by Molecular Dynamics (MD) simulations and Gaussian Network Model (GNM). The interplay of the inter-domain hinge with the cAMP and DNA binding domains are pre-disposed in the apo state as a conformational switch in the CRP's allosteric communication mechanism. The hinge at L134-D138 displaying intra- and inter-subunit coupled fluctuations with the cAMP and DNA binding domains leads to the emergence of stronger coupled fluctuations between the two domains and describes an on state. The flexible regions at K52-E58, P154/D155 and I175 maintain the dynamic coupling of the two domains. With a shift in the inter-domain hinge position towards the N terminus, nevertheless, the latter correlations between the domains loosen and become disordered; L134-D138 dynamically interacts only with the cAMP and DNA binding domains of its own subunit, and an off state is assumed. We present a mechanistic view on how the structural dynamic units are hierarchically built for the allosteric functional mechanism; from apo CRP monomer to apo-to-holo CRP dimers.

Highlights

The Escherichia coli cAMP Receptor Protein (CRP) activates the transcription of more than 150 genes
The Gaussian Network Model (GNM) cross-correlation maps of apo NMR structures, holo CRP crystal structure and the conformations from the apo CRP monomer/dimer and holo CRP dimer Molecular Dynamics (MD) simulations suggest a conformational switch mechanism mediated by the L134-D138 hinge in the allosteric communication of the DNA and cAMP binding domains
When we look at the dynamics of the unbound monomer, the elements of a plausible allosteric mechanism is still observed: The fluctuations of the position of the L134-D138 hinge to N terminus, the unstable correlated fluctuations of L134-D138 with K52-E58, and unstable correlated fluctuations between the DNA binding (P154-A156 and G173V176) and cAMP binding domains

Summary

Introduction

The Escherichia coli cAMP Receptor Protein (CRP) ( known as Catabolite Activator Protein, CAP) activates the transcription of more than 150 genes. The N-terminal domain, extending from V1 to N133, contains the primary cAMP binding site (anti cAMP) and mediates subunit-subunit interactions. This domain is formed by a helices A, B, C and eight b strands 1 to 8. The C-terminal domain extends from V139 to R209 with a helices D, E, F and four b strands 9 to 12, which has the helix–turn–helix (HTH) motif involved in the specific DNA and secondary cAMP binding (syn cAMP). To the HTH motif, residues A156-Q164 (activating region 1, AR1) are responsible for the transcription activation of lac class I and class II CRP-dependent promoters. In class II promoters, the interaction with RNAp is complemented by two more activating regions of residues H19, H21, E96, and K101

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Conclusions

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