Publications by authors named "Christopher Peralta"

3 Publications

  • Page 1 of 1

Structure of the FA core ubiquitin ligase closing the ID clamp on DNA.

Nat Struct Mol Biol 2021 Mar 8;28(3):300-309. Epub 2021 Mar 8.

Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA.

The Fanconi anemia (FA) pathway is essential for the repair of DNA interstrand crosslinks. Central to the pathway is the FA core complex, a ubiquitin ligase of nine subunits that monoubiquitinates the FANCI-FANCD2 (ID) DNA clamp. The 3.1 Å structure of the 1.1-MDa human FA core complex, described here, reveals an asymmetric assembly with two copies of all but the FANCC, FANCE and FANCF subunits. The asymmetry is crucial, as it prevents the binding of a second FANCC-FANCE-FANCF subcomplex that inhibits the recruitment of the UBE2T ubiquitin conjugating enzyme, and instead creates an ID binding site. A single active site then ubiquitinates FANCD2 and FANCI sequentially. We also present the 4.2-Å structures of the human core-UBE2T-ID-DNA complex in three conformations captured during monoubiquitination. They reveal the core-UBE2T complex remodeling the ID-DNA complex, closing the clamp on the DNA before ubiquitination. Monoubiquitination then prevents clamp opening after release from the core.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41594-021-00568-8DOI Listing
March 2021

DNA clamp function of the monoubiquitinated Fanconi anaemia ID complex.

Nature 2020 04 11;580(7802):278-282. Epub 2020 Mar 11.

Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY, USA.

The ID complex, involving the proteins FANCI and FANCD2, is required for the repair of DNA interstrand crosslinks (ICL) and related lesions. These proteins are mutated in Fanconi anaemia, a disease in which patients are predisposed to cancer. The Fanconi anaemia pathway of ICL repair is activated when a replication fork stalls at an ICL; this triggers monoubiquitination of the ID complex, in which one ubiquitin molecule is conjugated to each of FANCI and FANCD2. Monoubiquitination of ID is essential for ICL repair by excision, translesion synthesis and homologous recombination; however, its function remains unknown. Here we report a cryo-electron microscopy structure of the monoubiquitinated human ID complex bound to DNA, and reveal that it forms a closed ring that encircles the DNA. By comparison with the structure of the non-ubiquitinated ID complex bound to ICL DNA-which we also report here-we show that monoubiquitination triggers a complete rearrangement of the open, trough-like ID structure through the ubiquitin of one protomer binding to the other protomer in a reciprocal fashion. These structures-together with biochemical data-indicate that the monoubiquitinated ID complex loses its preference for ICL and related branched DNA structures, and becomes a sliding DNA clamp that can coordinate the subsequent repair reactions. Our findings also reveal how monoubiquitination in general can induce an alternative protein structure with a new function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41586-020-2110-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7398534PMC
April 2020

Structure of the cell-binding component of the binary toxin reveals a di-heptamer macromolecular assembly.

Proc Natl Acad Sci U S A 2020 01 2;117(2):1049-1058. Epub 2020 Jan 2.

Department of Biochemistry & Molecular Biology, University of Maryland School of Medicine, University of Maryland, Baltimore, MD 21201;

Targeting infection is challenging because treatment options are limited, and high recurrence rates are common. One reason for this is that hypervirulent strains often have a binary toxin termed the toxin, in addition to the enterotoxins TsdA and TsdB. The toxin has an enzymatic component, termed CDTa, and a pore-forming or delivery subunit termed CDTb. CDTb was characterized here using a combination of single-particle cryoelectron microscopy, X-ray crystallography, NMR, and other biophysical methods. In the absence of CDTa, 2 di-heptamer structures for activated CDTb (1.0 MDa) were solved at atomic resolution, including a symmetric (CDTb; 3.14 Å) and an asymmetric form (CDTb; 2.84 Å). Roles played by 2 receptor-binding domains of activated CDTb were of particular interest since the receptor-binding domain 1 lacks sequence homology to any other known toxin, and the receptor-binding domain 2 is completely absent in other well-studied heptameric toxins (i.e., anthrax). For CDTb, a Ca binding site was discovered in the first receptor-binding domain that is important for its stability, and the second receptor-binding domain was found to be critical for host cell toxicity and the di-heptamer fold for both forms of activated CDTb. Together, these studies represent a starting point for developing structure-based drug-design strategies to target the most severe strains of .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1919490117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6969506PMC
January 2020