Essential but sparse collagen hydroxylysyl post-translational modifications detected by DNP NMR

Abstract

COMMUNICATION Hartmut Oschkinat, Melinda J. Duer et al. Essential but sparse collagen hydroxylysyl post-translational modifications detected by DNP NMR The sparse but functionally essential post-translational collagen modification 5-hydroxylysine can undergo further transformations, including crosslinking, O-glycosylation, and glycation. Dynamic nuclear polarization (DNP) and stable isotope enriched lysine incorporation provide sufficient solid-state NMR sensitivity to identify these adducts directly in skin and vascular smooth muscle cell extracellular matrix (ECM), without extraction procedures, by comparison with chemical shifts of model compounds. Thus, DNP provides access to the elucidation of structural consequences of collagen modifications in intact tissue.

Design Parameters of Tissue‐Engineering Scaffolds at the Atomic Scale

“…This is a particularly noteworthy finding, given that collagen, the major fibrillar protein of the natural extracellular matrix, has a high proline content (approximately 10 %) yet has only moderate adhesion and very low differentiation potential for hNSCs in vitro . Interestingly, pioneering solid‐state NMR studies of bone collagen and cartilage show that prolines are involved in expansion joints that confer flexibility to collagen fibers, which is vital for the functional mechanics of these native tissues. This finding would suggest that different cell types require different material properties at the atomic‐length scale, opening the door to future investigations of novel biomaterials dedicated for other targeted tissues.…”

Design Parameters of Tissue‐Engineering Scaffolds at the Atomic Scale

“…Moreover,wedemonstrate that the molecular homogeneity of the scaffold relates to favorable functional properties.T he order of the stem-cell scaffold presumably enables an optimal exposure of the functional BMHP1 motif that fosters beneficial interactions with hNSCs, [20] while the exposure of the BMHP1 motif is suboptimal in disordered scaffolds due to ad istribution of favorable and unfavorable conformations.Inthe functionally most beneficial hydrogels, homogeneity is achieved by the removal of prolines from the BMHP1 motif.This is aparticularly noteworthy finding, given that collagen, the major fibrillar protein of the natural extracellular matrix, has ah igh proline content (approximately 10 %) [46] yet has only moderate adhesion and very low differentiation potential for hNSCs in vitro. [47] Interestingly, pioneering solid-state NMR studies of bone collagen and cartilage [46,[48][49][50][51][52][53][54][55][56] show that prolines are involved in expansion joints that confer flexibility to collagen fibers, [57] which is vital for the functional mechanics of these native tissues.T his finding would suggest that different cell types require different material properties at the atomic-length scale,opening the door to future investigations of novel biomaterials dedicated for other targeted tissues.M oreover,w hile it is well known that the mechanical properties of stem-cell scaffolds correlate with the functional performance of biological and designer matrices, [5] we show here that the rigidity at the molecular level also correlates with function. This is an important finding given that macroscopic characterizations of material properties only provide an average over atomistic, nanoscopic,and microscopic scales.Wealso like to underscore that previous microscopic biophysical characterizations did not succeed in quantifying the molecular differences of the stemcell scaffolds discussed in this study.…”

Targetable Tetrazine‐Based Dynamic Nuclear Polarization Agents for Biological Systems

“…Dynamic nuclear polarization (DNP) has had a significant impact on solid‐state nuclear magnetic resonance investigations of complex biological systems . With now routine signal enhancements in the 100‐fold range, DNP has aided the in vitro structural studies of amyloid polymers, membrane proteins, and biological material such as collagen, bones and tissues . Improvements in DNP instrumentation, sample preparation, and polarization agent design have also allowed NMR spectroscopists to turn their attention to the cellular environment, where the structural analysis of endogenous concentrations of biological macromolecules has come within reach .…”