Soluble expression of human glycoprotein Ibα in Escherichia coli through replacement of the N-terminal capping domain

Ryou, Jeong-Hyun; Park, Keunwan; Lee, Joong-Jae; Kim, Dongsup; Kim, Kwang Ho

doi:10.1016/j.pep.2014.06.001

Cited by 4 publications

(4 citation statements)

References 18 publications

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“…To improve the expression of recombinant OMD in E. coli , we employed a construct composed of a large portion of OMD and the N-terminal region of the leucine-rich repeat domain of internalin B (InlB), an approach previously reported for other proteins 20 . The chimeric OMD (chOMD) was designed by structural homology between OMD and InlB (Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

Molecular basis for governing the morphology of type-I collagen fibrils by Osteomodulin

et al. 2018

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Small leucine-rich repeat proteoglycan (SLRP) proteins have an important role in the organization of the extracellular matrix, especially in the formation of collagen fibrils. However, the mechanism governing the shape of collagen fibrils is poorly understood. Here, we report that the protein Osteomodulin (OMD) of the SLRP family is a monomeric protein in solution that interacts with type-I collagen. This interaction is dominated by weak electrostatic forces employing negatively charged residues of OMD, in particular Glu284 and Glu303, and controlled by entropic factors. The protein OMD establishes a fast-binding equilibrium with collagen, where OMD may engage not only with individual collagen molecules, but also with the growing fibrils. This weak electrostatic interaction is carefully balanced so it modulates the shape of the fibrils without compromising their viability.

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Section: Resultsmentioning

confidence: 99%

Molecular basis for governing the morphology of type-I collagen fibrils by Osteomodulin

et al. 2018

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“…Genes were synthesized for proteins containing 5 to 7 idealized building block modules. The N–terminal capping domain of internalin B was fused to DLRR_A and DLRR_B to enhance protein solubility and expression 12 , 20 whereas DLRR_C was expressed without a capping motif; instead the sequences of the N and C terminal repeats were redesigned to eliminate exposed hydrophobic residues. The idealized repeat designs were expressed in E.coli and found to be soluble and to have high thermal stability ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…For example, Toll-like receptor 4 (TLR4) contains three distinct regions of LRR repeats, each having different curvatures which collectively generate a surface with high shape complementarity to the target surface of the MD2 protein 10 . Current engineering approaches have focused on changing residues at the binding surfaces of an already existing or consensus repeat protein 11 - 16 , varying the numbers of repeat modules 17 - 19 , and fusing naturally occurring repeat proteins 10 , 20 , 21 . While powerful, these strategies do not allow customization of repeat protein curvature for a specific application.…”

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confidence: 99%

Control of repeat-protein curvature by computational protein design

Park

Shen

Parmeggiani

et al. 2015

Nat Struct Mol Biol

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Shape complementarity is an important component of molecular recognition, and the ability to precisely adjust the shape of a binding scaffold to match a target of interest would greatly facilitate the creation of high affinity protein reagents and therapeutics. Here we describe a general approach to control the shape of the binding surface on repeat protein scaffolds, and apply it to leucine rich repeat proteins. First, a set of self-compatible building block modules are designed that when polymerized each generate surfaces with unique but constant curvatures. Second, a set of junction modules that connect the different building blocks are designed. Finally, new proteins with custom designed shapes are generated by appropriately combining building block and junction modules. Crystal structures of the designs illustrate the power of the approach in controlling repeat protein curvature.

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“…Gene coding for the B‐subunit was obtained from the genomic DNA of E. coli O157:H7 (KCCM, Korea), and the other genes were synthesized (Bioneer, Korea). The cloning, expression, and purification of the proteins were carried out as described in our previous work (Ryou et al, ). Briefly, the gene coding for StxB‐TDP‐EGFP was cloned into a pET21a vector.…”

Section: Methodsmentioning

confidence: 99%

Engineering of bacterial exotoxins for highly efficient and receptor‐specific intracellular delivery of diverse cargos

Ryou

Sohn

Hwang

et al. 2016

Biotech & Bioengineering

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The intracellular delivery of proteins with high efficiency in a receptor-specific manner is of great significance in molecular medicine and biotechnology, but remains a challenge. Herein, we present the development of a highly efficient and receptor-specific delivery platform for protein cargos by combining the receptor binding domain of Escherichia coli Shiga-like toxin and the translocation domain of Pseudomonas aeruginosa exotoxin A. We demonstrated the utility and efficiency of the delivery platform by showing a cytosolic delivery of diverse proteins both in vitro and in vivo in a receptor-specific manner. In particular, the delivery system was shown to be effective for targeting an intracellular protein and consequently suppressing the tumor growth in xenograft mice. The present platform can be widely used for intracellular delivery of diverse functional macromolecules with high efficiency in a receptor-specific manner. Biotechnol. Bioeng. 2016;113: 1639-1646. © 2016 Wiley Periodicals, Inc.

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Soluble expression of human glycoprotein Ibα in Escherichia coli through replacement of the N-terminal capping domain

Cited by 4 publications

References 18 publications

Molecular basis for governing the morphology of type-I collagen fibrils by Osteomodulin

Molecular basis for governing the morphology of type-I collagen fibrils by Osteomodulin

Control of repeat-protein curvature by computational protein design

Engineering of bacterial exotoxins for highly efficient and receptor‐specific intracellular delivery of diverse cargos

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