A mini-review on the enzyme-mediated manipulation of proteins/peptides

Lin, Shaomin; He, Chunmao

doi:10.1016/j.cclet.2018.05.006

Cited by 14 publications

(8 citation statements)

References 117 publications

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“…Surface modification of the substrate is one of the key objectives in biosensor design to unite the biorecognition chemistry with the measurement system and integrate the core analytical reagent [22]. This could be replaced by posttranslational addition of specific peptides [23,24]. Nevertheless, taking a synthetic biology approach to the biorecognition reagent, suggests that a modular synthetic protein design, able to perform the tasks of both biorecognition and immobilization could eliminate a number of steps in the production pathway.…”

Section: Introductionmentioning

confidence: 99%

Gene to diagnostic: Self immobilizing protein for silica microparticle biosensor, modelled with sarcosine oxidase

et al. 2019

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A rational design approach is proposed for a multifunctional enzyme reagent for point-of-care diagnostics. The biomaterial reduces downstream isolation steps and eliminates immobilization coupling chemicals for integration in a diagnostic platform. Fusion constructs combined the central functional assay protein (e.g. monomeric sarcosine oxidase, mSOx, horseradish peroxidase, HRP), a visualizing protein (e.g. mCherry) and an in-built immobilization peptide (e.g. R5). Monitoring protein expression in E.coli was facilitated by following the increase in mCherry fluorescence, which could be matched to a color card, indicating when good protein expression has occurred. The R5 peptide (SSKKSGSYSGSKGSKRRIL) provided inbuilt affinity for silica and an immobilization capability for a silica based diagnostic, without requiring additional chemical coupling reagents. Silica particles extracted from beach sand were used to collect protein from crude protein extract with 85-95% selective uptake. The silica immobilized R5 proteins were stable for more than 2 months at room temperature. The Km for the silica-R52-mCh-mSOx-R5-6H was 16.5±0.9mM (compared with 16.5±0.4 mM, 16.3±0.3 mM, and 16.1±0.4 mM for R52-mCh-mSOx-R5-6H, mSOx-R5-6H and mSOx-6H respectively in solution). The use of the "silica-enzymes" in sarcosine and peroxide assays was shown, and a design using particle sedimentation through the sample was examined. Using shadowgraphy and particle image velocimetry the particle trajectory through the sample was mapped and an hourglass design with a narrow waist shown to give good control of particle position. The hourglass biosensor was demonstrated for sarcosine assay in the clinically useful range of 2.5 to 10 µM in both a dynamic and end point measurement regime.

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Section: Introductionmentioning

confidence: 99%

Gene to diagnostic: Self immobilizing protein for silica microparticle biosensor, modelled with sarcosine oxidase

et al. 2019

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“…Excellent articles summarized the different sortase classes with their functions in nature and structures, 2,4 mechanistic studies, 3,4 sortase inhibitors, 5,6 pili construction, 76,77 peptide and protein cyclisation, 63,78 general application elds and the use for protein engineering, [79][80][81][82][83][84][85] as well as the improvement of SML technology 11,12 and its differentiation with other enzymatic methods. [86][87][88][89][90] In this review article, we give an overview about the application of sortases in different interdisciplinary research elds. Using sortase A, a protein can be ligated to another biomolecule, a small synthetic molecule, a polymer or a surface.…”

Section: Introductionmentioning

confidence: 99%

Broadening the scope of sortagging

2019

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“…It was engineered by converting catalytic Ser221 to Cys, thereby increasing the ligase activity compared to amidase activity, and Residue Pro225 was converted to Ala to reduce steric assembling (Haridas et al, 2014). Subtiligase facilitates the ligation between a peptide C-terminal ester and a peptide N-terminal α-amine, without requiring a recognition motif at the termini of any reaction partners (Lin and He, 2018) (Figure 3C). The selective modification of the α-amine using subtiligase is a powerful approach in proteomics to enrich new N-termini arising from protease recognition and cleavage (Wiita et al, 2014), because 80% and 90% of wild-type eukaryotic proteins are acetylated at the N-terminal position (Polevoda and Sherman, 2003).…”

Section: Subtiligasementioning

confidence: 99%

“…In spite of being C-terminal-specific for Asx, this enzyme accepts most N-terminal amino acids to mediate intermolecular peptide and protein ligation (Nguyen et al, 2016). Although it was recently discovered, butelase 1 has been used for several purposes, such as protein modification and engineering, peptide/ protein ligation and labeling, peptide/protein macrocyclization, and living-cell surface labeling (Lin and He, 2018). No work has yet reported butelase 1 as being used for PEGylation reactions.…”

Section: Butelasementioning

confidence: 99%

From Synthesis to Characterization of Site-Selective PEGylated Proteins

et al. 2019

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Covalent attachment of therapeutic proteins to polyethylene glycol (PEG) is widely used for the improvement of its pharmacokinetic and pharmacological properties, as well as the reduction in reactogenicity and related side effects. This technique named PEGylation has been successfully employed in several approved drugs to treat various diseases, even cancer. Some methods have been developed to obtain PEGylated proteins, both in multiple protein sites or in a selected amino acid residue. This review focuses mainly on traditional and novel examples of chemical and enzymatic methods for site-selective PEGylation, emphasizing in N-terminal PEGylation, that make it possible to obtain products with a high degree of homogeneity and preserve bioactivity. In addition, the main assay methods that can be applied for the characterization of PEGylated molecules in complex biological samples are also summarized in this paper.

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A mini-review on the enzyme-mediated manipulation of proteins/peptides

Cited by 14 publications

References 117 publications

Gene to diagnostic: Self immobilizing protein for silica microparticle biosensor, modelled with sarcosine oxidase

Gene to diagnostic: Self immobilizing protein for silica microparticle biosensor, modelled with sarcosine oxidase

Broadening the scope of sortagging

From Synthesis to Characterization of Site-Selective PEGylated Proteins

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