Poster abstracts
Poster number 8 submitted by Tyler Billings
A secondary structure transition stabilizes an auto-inhibited conformation in the apo Cre catalytic domain
Tyler D Billings (Ohio State Biochemistry Program), Aparna Unnikrishnan (Biophysics Graduate Program), Deepak Kumar Yadav (Department of Chemistry and Biochemistry), Jonathan Montgomery (Ohio State Biochemistry Program), Mark P. Foster (Department of Chemistry and Biochemistry)
Abstract:
Cre is a conservative site-specific tyrosine recombinase that exchanges genetic material between homologous loxP DNA sequences. Its 343 amino acids comprise two domains: an N-terminal “core-biding” domain (residues 1-126), and a C-terminal "catalytic” domain (residues 138-343), connected by an extended 11-residue linker. Cre is best known for its role in producing mouse models to study the progression and treatment of human disease. Site-specific recombination is achieved via a series of single-strand phosphoryl transfer reactions involving a tyrosine nucleophile and the scissile phosphate on each strand. Exchange of strands is orchestrated by a network of interfacial contacts between protomers in synaptic complexes involving two DNA strands and four Cre protomers. One critical set of contacts involves the nine C-terminal residues of each of the four protomers, which in synaptic complexes adopt an α-helical conformation and dock into adjacent protomers in trans.1 Spectroscopic and biochemical data have shown that, in the absence of DNA, the C-terminal residues dock in cis over the DNA binding and active site residues.2 The presence of a cis-docked state implies an auto-inhibition process that is expected to affect Cre’s DNA binding equilibrium and cleavage activity. We present evidence that the cis-docked state of the Cre catalytic domain undergoes a secondary structure transition between the α-helical conformation observed in tetrameric synaptic complexes and an intramolecular β-sheet formed between the C-terminal sequence and the linker. This remarkable secondary structure transition parallels other less dramatic examples of auto-inhibition in the tyrosine recombinase class of enzymes.
References:
1. Ghosh, Kaushik, et al. “Synapsis of loxP Sites by Cre Recombinase.” Journal of Biological Chemistry, vol. 282, no. 33, Aug. 2007, pp. 24004–16. DOI.org (Crossref), https://doi.org/10.1074/jbc.M703283200.
2. Unnikrishnan, Aparna, et al. “DNA Binding Induces a Cis -to- Trans Switch in Cre Recombinase to Enable Intasome Assembly.” Proceedings of the National Academy of Sciences, vol. 117, no. 40, Oct. 2020, pp. 24849–58. DOI.org (Crossref), https://doi.org/10.1073/pnas.2011448117.
Keywords: auto-inhibition, recombinase, NMR