7W4S

Structure of the M. tuberculosis HtrA S363A mutant at room-temperature

Summary for 7W4S

• Entry DOI: 10.2210/pdb7w4s/pdb
• Descriptor: Probable serine protease HtrA1 (2 entities in total)
• Functional Keywords: htra family of serine protease, chymotrypsin-like, periplasm, protein quality control, signal transduction and room-temperature data., hydrolase
• Biological source: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
• Total number of polymer chains: 1
• Total formula weight: 32506.39

Authors

Gupta, A.K.,Gopal, B. (deposition date: 2021-11-29, release date: 2022-12-07, Last modification date: 2023-11-29)

Primary citation

Gupta, A.K.,Singh, K.,Patidar, Y.,Sharma, R.,Sardesai, A.A.,Reddy, G.,Gopal, B.
Allosteric Determinants in High Temperature Requirement A Enzymes Are Conserved and Regulate the Population of Active Conformations.
Acs Chem.Biol., 18:1487-1499, 2023

PubMed Abstract: High temperature requirement A (HtrA) are allosterically regulated enzymes wherein effector binding to the PDZ domain triggers proteolytic activity. Yet, it remains unclear if the inter-residue network governing allostery is conserved across HtrA enzymes. Here, we investigated and identified the inter-residue interaction networks by molecular dynamics simulations on representative HtrA proteases, DegS and PepD, in effector-bound and free forms. This information was used to engineer mutations that could potentially perturb allostery and conformational sampling in a different homologue, HtrA. Mutations in HtrA perturbed allosteric regulation─a finding consistent with the hypothesis that the inter-residue interaction network is conserved across HtrA enzymes. Electron density from data collected on cryo-protected HtrA crystals revealed that mutations altered the topology of the active site. Ensemble models fitted into electron density calculated from room-temperature diffraction data showed that only a fraction of these models had a catalytically competent active site conformation alongside a functional oxyanion hole thus providing experimental evidence that these mutations influenced conformational sampling. Mutations at analogous positions in the catalytic domain of DegS perturbed the coupling between effector binding and proteolytic activity, thus confirming the role of these residues in the allosteric response. The finding that a perturbation in the conserved inter-residue network alters conformational sampling and the allosteric response suggests that an ensemble allosteric model best describes regulated proteolysis in HtrA enzymes.

PubMed: 37319329
DOI: 10.1021/acschembio.2c00921

Experimental method

X-RAY DIFFRACTION (2.6 Å)