Pro-metastatic collagen lysyl hydroxylase dimer assemblies stabilized by Fe2+-binding

Abstract

Collagen lysyl hydroxylases (LH1-3) are Fe2+- and 2-oxoglutarate (2-OG)-dependent oxygenases that maintain extracellular matrix homeostasis. High LH2 levels cause stable collagen cross-link accumulations that promote fibrosis and cancer progression. However, developing LH antagonists will require structural insights. Here, we report a 2 Å crystal structure and X-ray scattering on dimer assemblies for the LH domain of L230 in Acanthamoeba polyphaga mimivirus. Loop residues in the double-stranded β-helix core generate a tail-to-tail dimer. A stabilizing hydrophobic leucine locks into an aromatic tyrosine-pocket on the opposite subunit. An active site triad coordinates Fe2+. The two active sites flank a deep surface cleft that suggest dimerization creates a collagen-binding site. Loss of Fe2+-binding disrupts the dimer. Dimer disruption and charge reversal in the cleft increase Km and reduce LH activity. Ectopic L230 expression in tumors promotes collagen cross-linking and metastasis. These insights suggest inhibitor targets for fibrosis and cancer.

SiMPLOD, a Structure-Integrated Database of Collagen Lysyl Hydroxylase (LH/PLOD) Enzyme Variants

“…As default option, the LH/PLOD dimer is colored using blue for the N‐terminal glycosyltransferase (GT) domain, orange for the central accessory (AC) domain, and green for the C‐terminal lysyl‐hydroxylase (LH) domain, as described, and the view can be re‐centered on the mutation(s) under investigation. A drop‐down menu allows to select the molecular structure for visualization, including the available LH3/PLOD3 and mimivirus L230 crystal structures, but also homology models of full‐length LH1/PLOD1 and LH2/PLOD2 enzymes. Mouse controls allow rotation (left button), translation (right button), zoom (wheel), and re‐centering (center button).…”

“…AC ¼ accessory domain; AKG ¼ 2-oxoglutarate; GT ¼ glycosyltransferase domain; LH ¼ lysyl-hydroxylase domain; UDP ¼ uridine diphosphate. menu allows to select the molecular structure for visualization, including the available LH3/PLOD3 (5) and mimivirus L230 (6) crystal structures, but also homology models of full-length LH1/ PLOD1 and LH2/PLOD2 enzymes. Mouse controls allow rotation (left button), translation (right button), zoom (wheel), and recentering (center button).…”

“…For macromolecular structures containing monomeric assemblies in the asymmetric unit (ie, human LH3 structures), dimeric ensembles were generated using COOT : molecular coordinates were loaded and crystallographic symmetry mates were displayed using the “show symmetry” option. Symmetry mates were manually inspected to identify the molecular partner of the monomer in the asymmetric unit to form physiological dimers, by looking at molecules forming crystal contacts near the dimerization interface defined by LH3 residues Arg714 and Leu715 . Coordinates of the dimer partner were saved using the “save symmetry coordinates” command.…”

“…Symmetry mates were manually inspected to identify the molecular partner of the monomer in the asymmetric unit to form physiological dimers, by looking at molecules forming crystal contacts near the dimerization interface defined by LH3 residues Arg714 (5) and Leu715. (6) Coordinates of the dimer partner were saved using the "save symmetry coordinates" command. The two monomers were combined together into a single PDB file using the "merge molecules" command in COOT.…”

Genotype-Phenotype Correlation of PLOD2 Skeletal Dysplasias Using Structural Information

“…Guo and colleagues have determined the 2Å crystal structure of the lysyl hydroxylase (LH) domain (amino acids 680–895 corresponding to human PLOD2 amino acids 548–758; Fig. ) of L230, an enzyme of Acanthamoeba polyphaga mimivirus (APMV) . They show that this domain confers metastatic potential to cancer cells.…”