Glycosylated amyloid‐like proteins in the structural extracellular polymers of aerobic granular sludge enriched with ammonium‐oxidizing bacteria

Lin, Yuemei; Reino, Clara; Carrera, Julián; Pérez, Julio; Loosdrecht, Mark C.M. van

doi:10.1002/mbo3.616

Cited by 63 publications

(29 citation statements)

References 64 publications

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“…We would expect the EPS to be similarly complex at a molecular level. Hence, full-scale systems may not be the ideal (Pronk et al 2017) and glycosylated proteins (Lin et al 2018). However, we still need to understand how widespread these EPS are in biofilms, as well as identify new extracellular polymers from other key systems to expand our database of identified, characterized and relevant EPS.…”

Section: Agreeing On Model Biofilms For Eps Characterizationmentioning

confidence: 99%

Extracellular polymeric substances of biofilms: Suffering from an identity crisis

et al. 2019

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Microbial biofilms can be both cause and cure to a range of emerging societal problems including antimicrobial tolerance, water sanitation, water scarcity and pollution. The identities of extracellular polymeric substances (EPS) responsible for the establishment and function of biofilms are poorly understood. The lack of information on the chemical and physical identities of EPS limits the potential to rationally engineer biofilm processes, and impedes progress within the water and wastewater sector towards a circular economy and resource recovery. Here, a multidisciplinary roadmap for addressing this EPS identity crisis is proposed. This involves improved EPS extraction and characterization methodologies, crossreferencing between model biofilms and full-scale biofilm systems, and functional description of isolated EPS with in situ techniques (e.g. microscopy) coupled with genomics, proteomics and glycomics. The current extraction and spectrophotometric characterization methods, often based on the principle not to compromise the integrity of the microbial cells, should be critically assessed, and more comprehensive methods for recovery and characterization of EPS need to be developed.

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Section: Agreeing On Model Biofilms For Eps Characterizationmentioning

confidence: 99%

Extracellular polymeric substances of biofilms: Suffering from an identity crisis

et al. 2019

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“…The contribution of glycosylation to amyloidogenic proteins is still a relatively unexplored field, although the effect of glycosylation of the Amyloid Precursor Protein of Aβ in Alzheimer's disease has gained attention due to its regulatory role in the protein's proteolytic processing (56,57). Bacterial amyloids derived from glycoproteins have now been reported as components of the structural extracellular matrix of biofilms rich in ammonia-oxidizing bacteria and nitrite-oxidizing bacteria grown in aerobic and granular sludge (58). Further characterization of glycoprotein-derived amyloids is warranted as these may demonstrate common features in different domains of life.…”

Section: Discussionmentioning

confidence: 99%

Amyloid aggregation of Streptococcus mutans Cnm influences its collagen-binding activity

Cologna

Samaddar²,

Valle

et al. 2021

Preprint

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The glycosylated collagen- and laminin-binding surface adhesin Cnm is present in approximately 20% of S. mutans clinical isolates and is associated with systemic infections and increased caries risk. Other surface-associated collagen-binding proteins of S. mutans such as P1 and WapA have been demonstrated to form an amyloid quaternary structure with functional implications within biofilms. In silico analysis predicted that the b-sheet rich N-terminal collagen-binding domain (CBD) of Cnm has propensity for amyloid aggregation, whereas the threonine-rich C-terminal domain was predicted to be disorganized. In this study, thioflavin-T fluorescence and electron microscopy were used to show that Cnm forms amyloids either in its native glycosylated or recombinant non-glycosylated forms and that the CBD of Cnm is the main amyloidogenic unit of Cnm. We then performed a series of in vitro, ex vivo and in vivo assays to characterize the amylogenic properties of Cnm. In addition, Congo red birefringence indicated that Cnm is a major amyloidogenic protein of S. mutans biofilms. Competitive binding assays using collagen-coated microtiter plates and dental roots, a substrate rich in collagen, revealed that Cnm monomers inhibit S. mutans binding to collagenous substrates whereas Cnm amyloid aggregates lose this property. Thus, while Cnm contributes to recognition and initial binding of S. mutans to collagen-rich surfaces, Cnm amyloid aggregation appears to represent a mechanism to modulate this activity in mature biofilms.

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“…However, DTAB-treated proteins were found to be incompatible with SDS-PAGE reagents due to smeared band distribution (Figure 3b). Specifically, secondary SDS hydrolysis and heating (100 °C) denatured the peptides and prevented protein aggregation, 60 which caused serious protein debris. 53 The low toxic and nonionic Tween 80 in combination with quartz sand met the requirement of MS-based detection (Figure 4).…”

Section: ■ Discussionmentioning

confidence: 99%

Development of a Quartz Sand Protocol for Exoproteome Exploration from Anammox Consortia

Chen

Huang

Meng

et al. 2020

ACS Sustainable Chem. Eng.

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A critical prerequisite for unbiased anaerobic ammonium oxidation (anammox) exoproteome exploration is the optimal extraction and treatment of extracellular polymeric substances (EPS). Herein, by leveraging the advantages of physical shear and potential electrostatic repulsion mechanisms, a nondestructive quartz sand protocol for in situ extraction of extracellular proteins (PN) for anammox consortia was developed and applied. According to an L16(45) orthogonal design, the optimal procedure, which contributes the maximum yield of EPS with minimal cell rupture, was identified as follows: 1.0 mm particle size, 100 g/g-VSS quartz sand ratio, 400 rpm, and 2 h. While achieving approximately the same yield of EPS, hierarchical clustering and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that the quartz sand extraction was highly similar to the conventional cation exchange resin (CER) treatment in protein secondary structure properties and protein band distribution patterns, compared to other methods. However, the number of identified proteins and hydrophobic protein proportions were significantly increased by 5.3-fold and 2.6-fold after quartz sand treatment (122, 11.5%), respectively, compared to that of the CER method (23, 4.4%), while low cell lysis (52.1% extracellular component) was obtained based on subcellular localization data. Strong correlations between hydrophobicity of method-specific proteins and β-sheet proportion (r 2 = 0.97, p = 0.0024), as well as architecture tightness (r 2 = 0.94, p = 0.0069) suggest that once the EPS protein structure becomes loose, exposed inner β-sheet residues are prone to express hydrophobic properties. Addition of surfactants exhibited an undesirable solubilization effect on the protein secondary structure, gel separation, and spatial proteome, irrespective of PN yields. Overall, this work demonstrates the feasibility and effectiveness of extracting anammox EPS using an in situ quartz method that can satisfy yield-lysis trade-offs, retain the natural EPS traits, and allow the collection of more extracellular proteomic information.

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Glycosylated amyloid‐like proteins in the structural extracellular polymers of aerobic granular sludge enriched with ammonium‐oxidizing bacteria

Cited by 63 publications

References 64 publications

Extracellular polymeric substances of biofilms: Suffering from an identity crisis

Extracellular polymeric substances of biofilms: Suffering from an identity crisis

Amyloid aggregation of Streptococcus mutans Cnm influences its collagen-binding activity

Development of a Quartz Sand Protocol for Exoproteome Exploration from Anammox Consortia

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