Oriented immobilization of proteins

Rao, Srivatsa V.; Anderson, Kimberly W.; Bachas, Leonidas G.

doi:10.1007/bf01243043

Cited by 242 publications

(187 citation statements)

References 118 publications

Supporting

Mentioning

181

Contrasting

Unclassified

Order By: Relevance

“…Addition of exogenous components to the native cellulosome has been proposed earlier in the form of a "supercellulosome" where exogenous enzymes are incorporated into the intact cellulosome using bifunctional crosslinking reagents (14,36); however, the nonspecific chemical nature of crosslinking could impair the activities of the β-glucosidase and the cellulosomal enzymes (37). In contrast, our data show that the use of fused CohII module allows specific incorporation of the enzyme into the cellulosome that preserves its essential cellobiase activity.…”

Section: Discussioncontrasting

confidence: 53%

Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Gefen

Anbar

Morag³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

111

View full text Add to dashboard Cite

The conversion of recalcitrant plant-derived cellulosic biomass into biofuels is dependent on highly efficient cellulase systems that produce near-quantitative levels of soluble saccharides. Similar to other fungal and bacterial cellulase systems, the multienzyme cellulosome system of the anaerobic, cellulolytic bacterium Clostridium thermocellum is strongly inhibited by the major end product cellobiose. Cellobiose-induced inhibition can be relieved via its cleavage to noninhibitory glucose by the addition of exogenous noncellulosomal enzyme β-glucosidase; however, because the cellulosome is adsorbed to the insoluble substrate only a fraction of β-glucosidase would be available to the cellulosome. Towards this end, we designed a chimeric cohesin-fused β-glucosidase (BglA-CohII) that binds directly to the cellulosome through an unoccupied dockerin module of its major scaffoldin subunit. The β-glucosidase activity is thus focused at the immediate site of cellobiose production by the cellulosomal enzymes. BglA-CohII was shown to retain cellobiase activity and was readily incorporated into the native cellulosome complex. Surprisingly, it was found that the native C. thermocellum cellulosome exists as a homooligomer and the high-affinity interaction of BglA-CohII with the scaffoldin moiety appears to dissociate the oligomeric state of the cellulosome. Complexation of the cellulosome and BglA-CohII resulted in higher overall degradation of microcrystalline cellulose and pretreated switchgrass compared to the native cellulosome alone or in combination with wild-type BglA in solution. These results demonstrate the effect of enzyme targeting and its potential for enhanced degradation of cellulosic biomass.

show abstract

Section: Discussioncontrasting

confidence: 53%

Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Gefen

Anbar

Morag³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

111

View full text Add to dashboard Cite

show abstract

“…[20][21][22]). Biomolecules are commonly bound to surfaces via amino, carboxyl, or thiol groups on the protein and/or the substrate.…”

Section: Discussionmentioning

confidence: 99%

Immobilization of glycoproteins, such as VEGF, on biodegradable substrates

Sharon

Puleo

2008

Acta Biomaterialia

View full text Add to dashboard Cite

Attachment of growth factors to biodegradable polymers, such as poly(lactide-co-glycolide) (PLGA), may enhance and/or accelerate integration of tissue engineering scaffolds. Although proteins are commonly bound via abundant amino groups, a more selective approach may increase bioactivity of immobilized molecules. In this research, exposed carboxyl groups on acid-terminated PLGA were modified with dihydrazide spacer molecules. The number of hydrazide groups available for subsequent attachment of protein was dependent on dihydrazide length, with shorter molecules present at significantly greater surface densities. The potent angiogenic glycoprotein vascular endothelial growth factor (VEGF) was oxidized with periodate and the aldehyde moieties allowed to react with the hydrazide-derivatized PLGA. Derivatization initially affected the amount of protein bound to the surfaces, but differences were substantially reduced following overnight incubation in saline. More importantly, use of shorter dihydrazide spacers significantly enhanced accessibility of immobilized VEGF for binding neutralizing antibody and soluble VEGR receptor. Furthermore, immobilized growth factor enhanced endothelial cell proliferation, with surfaces having the shortest and longest spacers stimulating greater effects. The present work has not only demonstrated an alternative approach to immobilizing growth factors on biodegradable materials, but the scheme can be used to alter the amount of protein bound as well as its availability for subsequent biointeractions.

show abstract

“…128,129 The incorporation of cysteine on the Ab fragment apart from the antigen recognition site enables the use of sulfhydryl linkers that leads to more efficient solidphase immobilization. 130,131 The cysteine tag also broadens the choice of solid supports as Abs can be immobilized on various substrates including cyanogen bromide (CNBr)-activated surfaces. 132 It provides the robustness to the tagged Ab required for leach-proof immobilization.…”

Section: Site-specificmentioning

confidence: 99%

Immobilization of Antibodies and Enzymes on 3-Aminopropyltriethoxysilane-Functionalized Bioanalytical Platforms for Biosensors and Diagnostics

et al. 2014

View full text Add to dashboard Cite

/npsi/ctrl?lang=en http://nparc.cisti-icist.nrc-cnrc.gc.ca/npsi/ctrl?lang=fr READ THESE TERMS AND CONDITIONS CAREFULLY BEFORE USING THIS WEBSITE.http://nparc.cisti-icist.nrc-cnrc.gc.ca/npsi/jsp/nparc_cp.jsp?lang=en Vous avez des questions? Nous pouvons vous aider. Pour communiquer directement avec un auteur, consultez la première page de la revue dans laquelle son article a été publié afin de trouver ses coordonnées. Si vous n'arrivez pas à les repérer, communiquez avec nous à PublicationsArchive-ArchivesPublications@nrc-cnrc.gc.ca.

show abstract

Oriented immobilization of proteins

Cited by 242 publications

References 118 publications

Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Enhanced cellulose degradation by targeted integration of a cohesin-fused β-glucosidase into the Clostridium thermocellum cellulosome

Immobilization of glycoproteins, such as VEGF, on biodegradable substrates

Immobilization of Antibodies and Enzymes on 3-Aminopropyltriethoxysilane-Functionalized Bioanalytical Platforms for Biosensors and Diagnostics

Contact Info

Product

Resources

About