Cloning, expression and purification of the SRCR domains of glycoprotein 340

Purushotham, Sangeetha; Deivanayagam, Champion

doi:10.1016/j.pep.2013.05.003

Cited by 6 publications

(2 citation statements)

References 29 publications

(34 reference statements)

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“…All gp340 domains have architectures that typically function in binding interactions, especially SRCR. Each SRCR domain, named for the order in which it appears beginning with SRCR1, is 100-110 residues, and there is a short flexible intervening segment between each, except the fourth and fifth domains (Purushotham and Deivanayagam, 2013). These connector segments are referred to as SIDs, and this is where the covalent attachment of O-glycans, which contribute to bacterial binding, occurs (Purushotham and Deivanayagam, 2014).…”

Section: Gp340 Structure and Interactionsmentioning

confidence: 99%

Proteins, Pathogens, and Failure at the Composite-Tooth Interface

et al. 2014

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In the United States, composites accounted for nearly 70% of the 173.2 million composite and amalgam restorations placed in 2006 (Kingman et al., 2012), and it is likely that the use of composite will continue to increase as dentists phase out dental amalgam. This trend is not, however, without consequences. The failure rate of composite restorations is double that of amalgam (Ferracane, 2013). Composite restorations accumulate more biofilm, experience more secondary decay, and require more frequent replacement. In vivo biodegradation of the adhesive bond at the composite-tooth interface is a major contributor to the cascade of events leading to restoration failure. Binding by proteins, particularly gp340, from the salivary pellicle leads to biofilm attachment, which accelerates degradation of the interfacial bond and demineralization of the tooth by recruiting the pioneer bacterium Streptococcus mutans to the surface. Bacterial production of lactic acid lowers the pH of the oral microenvironment, erodes hydroxyapatite in enamel and dentin, and promotes hydrolysis of the adhesive. Secreted esterases further hydrolyze the adhesive polymer, exposing the soft underlying collagenous dentinal matrix and allowing further infiltration by the pathogenic biofilm. Manifold approaches are being pursued to increase the longevity of composite dental restorations based on the major contributing factors responsible for degradation. The key material and biological components and the interactions involved in the destructive processes, including recent advances in understanding the structural and molecular basis of biofilm recruitment, are described in this review. Innovative strategies to mitigate these pathogenic effects and slow deterioration are discussed.

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Section: Gp340 Structure and Interactionsmentioning

confidence: 99%

Proteins, Pathogens, and Failure at the Composite-Tooth Interface

et al. 2014

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“…We studied the adherence of GtfB-CD to SRCR 1 by surface plasmon resonance (SPR) using a Biacore T200 instrument in the Multidisciplinary Molecular Interaction Core of the University of Alabama at Birmingham. The recombinant scavenger receptor cysteine-rich domain 1 expressed in insect cells (iSRCR 1 ) was immobilized on a carboxymethyl dextran chip (CM5) using an amine-coupling procedure as described previously (Mieher et al, 2018;Purushotham & Deivanayagam, 2013, 2014. Briefly, the chip surface was activated using N-hydroxysuccinimide and N-ethyl-N 0 -(3-diethylaminopropyl)carbodiimide, and the ligand (at 50 mg ml À1 ) in 10 mM sodium acetate buffer pH 4.0 was injected at a flow rate of 5 ml min À1 until the desired resonance units were obtained.…”

Section: Adherence Of Gtfb-cd To Srcr 1 Using Surface Plasmon Resonancementioning

confidence: 99%

The catalytic domains of Streptococcus mutans glucosyltransferases: a structural analysis

Schormann¹,

Patel²,

Thannickal³

et al. 2023

Acta Crystallogr F Struct Biol Commun

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Streptococcus mutans, found in the human oral cavity, is a significant contributor to the pathogenesis of dental caries. This bacterium expresses three genetically distinct types of glucosyltransferases named GtfB (GTF-I), GtfC (GTF-SI) and GtfD (GTF-S) that play critical roles in the development of dental plaque. The catalytic domains of GtfB, GtfC and GtfD contain conserved active-site residues for the overall enzymatic activity that relate to hydrolytic glycosidic cleavage of sucrose to glucose and fructose, release of fructose and generation of a glycosyl-enzyme intermediate in the reducing end. In a subsequent transglycosylation step, the glucosyl moiety is transferred to the nonreducing end of an acceptor to form a growing glucan polymer chain made up of glucose molecules. It has been proposed that both sucrose breakdown and glucan synthesis occur in the same active site of the catalytic domain, although the active site does not appear to be large enough to accommodate both functions. These three enzymes belong to glycoside hydrolase family 70 (GH70), which shows homology to glycoside hydrolase family 13 (GH13). GtfC synthesizes both soluble and insoluble glucans (α-1,3 and α-1,6 glycosidic linkages), while GtfB and GtfD synthesize only insoluble or soluble glucans, respectively. Here, crystal structures of the catalytic domains of GtfB and GtfD are reported. These structures are compared with previously determined structures of the catalytic domain of GtfC. With this work, apo structures and inhibitor-complex structures with acarbose are now available for the catalytic domains of GtfC and GtfB. The structure of GtfC with maltose allows further identification and comparison of active-site residues. A model of sucrose binding to GtfB is also included. The new structure of the catalytic domain of GtfD affords a structural comparison of the three S. mutans glycosyltransferases. Unfortunately, the catalytic domain of GtfD is not complete since crystallization resulted in the structure of a truncated protein lacking approximately 200 N-terminal residues of domain IV.

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The Calcium-induced Conformation and Glycosylation of Scavenger-rich Cysteine Repeat (SRCR) Domains of Glycoprotein 340 Influence the High Affinity Interaction with Antigen I/II Homologs

Purushotham

Deivanayagam

2014

Journal of Biological Chemistry

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Background: AgI/II homolog interaction with GP340 is crucial for bacterial attachment to tooth surface. Results: Tandem SRCR domains efficiently adhere/aggregate bacteria. Calcium-induced conformational switch and O-linked carbohydrates of SRCRs are necessary for the interaction with AgI/II homologs. Conclusion: High affinity interactions are dictated by calcium and carbohydrates. Significance: Oral streptococci adhere to specific calcium-induced conformation of immobilized SRCRs and to its carbohydrates.

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Cloning, expression and purification of the SRCR domains of glycoprotein 340

Cited by 6 publications

References 29 publications

Proteins, Pathogens, and Failure at the Composite-Tooth Interface

Proteins, Pathogens, and Failure at the Composite-Tooth Interface

The catalytic domains of Streptococcus mutans glucosyltransferases: a structural analysis

The Calcium-induced Conformation and Glycosylation of Scavenger-rich Cysteine Repeat (SRCR) Domains of Glycoprotein 340 Influence the High Affinity Interaction with Antigen I/II Homologs

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