biostudies-literatureGiladi MIsrael Cancer Research Fund (Israel Cancer Research Fund, Inc.)Israel Science FoundationIsrael Science Foundation (ISF)Israel Cancer Research Fund10786https://www.ebi.ac.uk/biostudies/studies/S-EPMC12663261biostudies-literatureUnknown16(1)Human cis-prenyltransferase (hcis-PT) synthesizes long-chain isoprenoids essential for N-linked protein glycosylation. This heteromeric complex comprises the catalytic subunit DHDDS and the regulatory Nogo-B receptor (NgBR). Although NgBR dramatically enhances DHDDS activity, the molecular basis for this allosteric regulation remains unclear. Here, we combined crystallography, hydrogen-deuterium exchange mass spectrometry (HDX-MS), molecular dynamics simulations, and network analysis to uncover the structural dynamics and communication pathways within hcis-PT. By solving the apo structure of hcis-PT, we reveal only a localized flexibility at the active site and the NgBR C-terminus. However, HDX-MS demonstrated widespread substrate-induced stabilization, particularly at the NgBR βD-βE loop,Nature communicationsStructural mechanisms of allosteric regulation in the human cis-prenyltransferase complex.PMC126632612337/251289067Man PGuardiani CHaitin YVankova PGiacomello AGiladi MAviram LKredi SGaizinger YfalseStructural mechanisms of allosteric regulation in the human cis-prenyltransferase complex.Human cis-prenyltransferase (hcis-PT) synthesizes long-chain isoprenoids essential for N-linked protein glycosylation. This heteromeric complex comprises the catalytic subunit DHDDS and the regulatory Nogo-B receptor (NgBR). Although NgBR dramatically enhances DHDDS activity, the molecular basis for this allosteric regulation remains unclear. Here, we combined crystallography, hydrogen-deuterium exchange mass spectrometry (HDX-MS), molecular dynamics simulations, and network analysis to uncover the structural dynamics and communication pathways within hcis-PT. By solving the apo structure of hcis-PT, we reveal only a localized flexibility at the active site and the NgBR C-terminus. However, HDX-MS demonstrated widespread substrate-induced stabilization, particularly at the NgBR βD-βE loop,2025-01-01T00:00:00Z2025 Nov2026-07-15T07:47:49.562Z2026-07-01T03:08:12.75ZS-EPMC126632614131534810.1038/s41467-025-65833-6