6W2H

Crystal Structure of the Internal UBA Domain of HHR23A

Summary for 6W2H

• Entry DOI: 10.2210/pdb6w2h/pdb
• Descriptor: UV excision repair protein RAD23 homolog A, SULFATE ION (3 entities in total)
• Functional Keywords: ubiquitin associated domain, uba domain, dna binding protein, helical bundle
• Biological source: Homo sapiens (Human)
• Total number of polymer chains: 1
• Total formula weight: 5736.43

Authors

Bowler, B.E.,Zeng, B.,Becht, D.C.,Rothfuss, M.,Sprang, S.R.,Mou, T.-C. (deposition date: 2020-03-05, release date: 2021-03-10, Last modification date: 2024-04-03)

Primary citation

Becht, D.C.,Leavens, M.J.,Zeng, B.,Rothfuss, M.T.,Briknarova, K.,Bowler, B.E.
Residual Structure in the Denatured State of the Fast-Folding UBA(1) Domain from the Human DNA Excision Repair Protein HHR23A.
Biochemistry, 61:767-784, 2022

PubMed Abstract: The structure of the first ubiquitin-associated domain from HHR23A, UBA(1), was determined by X-ray crystallography at a 1.60 Å resolution, and its stability, folding kinetics, and residual structure under denaturing conditions have been investigated. The concentration dependence of thermal denaturation and size-exclusion chromatography indicate that UBA(1) is monomeric. Guanidine hydrochloride (GdnHCl) denaturation experiments reveal that the unfolding free energy, Δ°'(HO), of UBA(1) is 2.4 kcal mol. Stopped-flow folding kinetics indicates sub-millisecond folding with only proline isomerization phases detectable at 25 °C. The full folding kinetics are observable at 4 °C, yielding a folding rate constant, , in the absence of a denaturant of 13,000 s and a Tanford β-value of 0.80, consistent with a compact transition state. Evaluation of the secondary structure via circular dichroism shows that the residual helical structure in the denatured state is replaced by polyproline II structure as the GdnHCl concentration increases. Analysis of NMR secondary chemical shifts for backbone NH, CO, and Cα atoms between 4 and 7 M GdnHCl shows three islands of residual helical secondary structure that align in sequence with the three native-state helices. Extrapolation of the NMR data to 0 M GdnHCl demonstrates that helical structure would populate to 17-33% in the denatured state under folding conditions. Comparison with NMR data for a peptide corresponding to helix 1 indicates that this helix is stabilized by transient tertiary interactions in the denatured state of UBA(1). The high helical content in the denatured state, which is enhanced by transient tertiary interactions, suggests a diffusion-collision folding mechanism.

PubMed: 35430812
DOI: 10.1021/acs.biochem.2c00011

Experimental method

X-RAY DIFFRACTION (1.6 Å)