Tuning cellular response by modular design of bioactive domains in collagen

Que, Richard; Chan, Sam Wei Polly; Jabaiah, Abeer; Lathrop, Richard H.; Silva, Nancy A. Da; Wang, Szu‐Wen

doi:10.1016/j.biomaterials.2015.02.074

Cited by 22 publications

(33 citation statements)

References 52 publications

(98 reference statements)

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“…Recombinant collagens have also been used to characterize and manipulate integrin binding. Recombinant human type III collagen expressed in yeast promoted cell binding; this binding could be eliminated through removal of the five known integrin binding sites and restored through insertion of GFOGER (36, 37). The insertion of the human type I collagen cell binding sequences GFPGER or GLPGER within the collagen domain of the bacterial collagen protein Scl2 resulted in integrin binding and cell adhesion (22, 29, 30), and insertion of distinctive sequences could be used to form hydrogels with selective adhesion to endothelial cells versus smooth muscle cells (38).…”

Section: Discussionmentioning

confidence: 99%

Mapping the Effect of Gly Mutations in Collagen on α2β1 Integrin Binding

Yiğit

An³

et al. 2016

Journal of Biological Chemistry

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The replacement of one Gly in the essential repeating tripeptide sequence of the type I collagen triple helix results in the dominant hereditary bone disorder osteogenesis imperfecta. The mechanism leading to pathology likely involves misfolding and autophagy, although it has been hypothesized that some mutations interfere with known collagen interactions. Here, the effect of Gly replacements within and nearby the integrin binding GFPGER sequence was investigated using a recombinant bacterial collagen system. When a six-triplet human type I collagen sequence containing GFPGER was introduced into a bacterial collagen-like protein, this chimeric protein bound to integrin. Constructs with Gly to Ser substitutions within and nearby the inserted human sequence still formed a trypsin-resistant triple helix, suggesting a small local conformational perturbation. Gly to Ser mutations within the two Gly residues in the essential GFPGER sequence prevented integrin binding and cell attachment as predicted from molecular dynamics studies of the complex. Replacement of Gly residues C-terminal to GFPGER did not affect integrin binding. In contrast, Gly replacements N-terminal to the GFPGER sequence, up to four triplets away, decreased integrin binding and cell adhesion. This pattern suggests either an involvement of the triplets N-terminal to GFPGER in initial binding or a propagation of the perturbation of the triple helix C-terminal to a mutation site. The asymmetry in biological consequences relative to the mutation site may relate to the observed pattern of osteogenesis imperfecta mutations near the integrin binding site.

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Section: Discussionmentioning

confidence: 99%

Mapping the Effect of Gly Mutations in Collagen on α2β1 Integrin Binding

Yiğit

An³

et al. 2016

Journal of Biological Chemistry

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“…The locations for introduction of IKVAV were chosen to limit the number of amino acids changed from the native sequence. For the creation of cysteine‐containing variants, the collagen III variant in which native binding motifs were replaced with non‐binding GSPGGK sequences (variant rCol‐0G), was utilized as a scaffold base . Introduced cysteine locations were chosen to avoid known integrin binding sites and the native MMP cleavage site.…”

Section: Methodsmentioning

confidence: 99%

“…To express our collagen variants, we utilized S. cerevisiae strain BYα2β2, which has two copies of human prolyl‐4‐hydroxylase inserted into its chromosomes . The full‐length collagen variant genes were introduced into CEN/ARS plasmids and transformed into BYα2β2.…”

Section: Methodsmentioning

confidence: 99%

“…The full‐length collagen variant genes were introduced into CEN/ARS plasmids and transformed into BYα2β2. Expression of collagen was induced, the yeast was harvested, and the resulting collagen variants were purified according to previously described protocols . Yeasts cells were mechanically disrupted using a Thermo Electron French Press and digested with pepsin.…”

Section: Methodsmentioning

confidence: 99%

“…We have previously demonstrated the ability to introduce the integrin binding motif GFOGER from collagen I into our non‐cell adhering variant of collagen III . However, the study was performed only in a two‐dimensional (2D) context.…”

Section: Introductionmentioning

confidence: 99%

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Tailoring Collagen to Engineer the Cellular Microenvironment

Que

Crakes

Abdulhadi

et al. 2018

Biotechnology Journal

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Collagen is the most abundant protein in the extracellular matrix (ECM), and it can direct the behavior of the neighboring cells. By customizing properties of collagen, it is possible to control the cells that interact with it. Utilizing a bottom-up strategy, modular gene fragments are assembled and recombinantly processed to create collagen-mimetic variants that modulate proteolytic degradation, cell adhesion, and mechanical characteristics. The removal of the native MMP cleavage site results in MMP-1 resistant collagen. By introducing additional MMP-susceptible sequences, the degradation characteristics of collagen molecules are modified. Additional non-native functionality is also introduced into the collagen, including the IKVAV sequence, which has been implicated in neurite outgrowth. This mutation, which disrupts the Gly-X-Y tripeptide repeat of collagen, does not prevent the formation of triple-helical collagen. Non-native cysteines and the integrin binding sequence GFOGER are combined in the collagen, and encapsulation of normal human lung fibroblasts within collagen hydrogels are tested. Cells remain spherical, when encapsulated within hydrogels of collagen variants in which the native integrin binding sites are removed, but cell adhesion is restored with the introduction of non-native GFOGER binding sequences. This modular collagen system allows for the combination of multiple functionalities, and it enables the production of biomimetic scaffolds with customizable characteristics to modulate cellular microenvironments.

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Incorporation of a Ligand Peptide for Immune Inhibitory Receptor LAIR‐1 on Biomaterial Surfaces Inhibits Macrophage Inflammatory Responses

Kim

Chu

Hsieh

et al. 2017

Adv Healthcare Materials

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Leukocyte-associated Ig-like receptor-1 (LAIR-1) is an inhibitory receptor broadly expressed on immune cells, with its ligands residing within the extracellular matrix protein collagen. In this study, we modify surfaces with a LAIR-1 ligand peptide (LP), and observe that macrophages cultured on LAIR-1 LP-conjugated surfaces exhibited significantly reduced secretion of inflammatory cytokines in response to proinflammatory stimuli that mimic an injured environment. These downregulated mediators include TNF-α, MIP-1α, MIP-1β, MIP-2, RANTES, and MIG. Knockdown of LAIR-1 using siRNA abrogates this inhibition of cytokine secretion, supporting the specificity of the inhibitory effect to this receptor. These results are the first to demonstrate that integration of LAIR-1 ligands with biomaterials could suppress inflammatory responses.

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Tuning cellular response by modular design of bioactive domains in collagen

Cited by 22 publications

References 52 publications

Mapping the Effect of Gly Mutations in Collagen on α2β1 Integrin Binding

Mapping the Effect of Gly Mutations in Collagen on α2β1 Integrin Binding

Tailoring Collagen to Engineer the Cellular Microenvironment

Incorporation of a Ligand Peptide for Immune Inhibitory Receptor LAIR‐1 on Biomaterial Surfaces Inhibits Macrophage Inflammatory Responses

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