77 research outputs found
Recommended from our members
Local orientational disorder in peptide fibrils probed by a combination of residue-specific 13C-18O labelling, polarised infrared spectroscopy and molecular combing
A novel combination of site-specific isotope labelling, polarised infrared spectroscopy and molecular combing reveal local orientational ordering in the fibril-forming peptide YTIAALLSPYSGGRADS. Use of 13C-18O labelled alanine residues demonstrates that the Nterminal end of the peptide is incorporated into the cross-beta structure, while the C-terminal end shows orientational disorde
The propensity of the bacterial rodlin protein RdlB to form amyloid fibrils determines its function in Streptomyces coelicolor.
Streptomyces bacteria form reproductive aerial hyphae that are covered with a pattern of pairwise aligned fibrils called rodlets. The presence of the rodlet layer requires two homologous rodlin proteins, RdlA and RdlB, and the functional amyloid chaplin proteins, ChpA-H. In contrast to the redundancy shared among the eight chaplins, both RdlA and RdlB are indispensable for the establishment of this rodlet structure. By using a comprehensive biophysical approach combined with in vivo characterization we found that RdlB, but not RdlA, readily assembles into amyloid fibrils. The marked difference in amyloid propensity between these highly similar proteins could be largely attributed to a difference in amino acid sequence at just three sites. Further, an engineered RdlA protein in which these three key amino acids were replaced with the corresponding residues from RdlB could compensate for loss of RdlB and restore formation of the surface-exposed amyloid layer in bacteria. Our data reveal that RdlB is a new functional amyloid and provide a biophysical basis for the functional differences between the two rodlin proteins. This study enhances our understanding of how rodlin proteins contribute to formation of an outer fibrillar layer during spore morphogenesis in streptomycetes
The Assembly of Individual Chaplin Peptides from Streptomyces coelicolor into Functional Amyloid Fibrils
The self-association of proteins into amyloid fibrils offers an alternative to the natively folded state of many polypeptides. Although commonly associated with disease, amyloid fibrils represent the natural functional state of some proteins, such as the chaplins from the soil-dwelling bacterium Streptomyces coelicolor, which coat the aerial mycelium and spores rendering them hydrophobic. We have undertaken a biophysical characterisation of the five short chaplin peptides ChpD-H to probe the mechanism by which these peptides self-assemble in solution to form fibrils. Each of the five chaplin peptides produced synthetically or isolated from the cell wall is individually surface-active and capable of forming fibrils under a range of solution conditions in vitro. These fibrils contain a highly similar cross-β core structure and a secondary structure that resembles fibrils formed in vivo on the spore and mycelium surface. They can also restore the growth of aerial hyphae to a chaplin mutant strain. We show that cysteine residues are not required for fibril formation in vitro and propose a role for the cysteine residues conserved in four of the five short chaplin peptides
Effect of rennet on the composition, proteolysis and microstructure of reduced-fat Cheddar cheese during ripening
Predictive models for upstream mammalian cell culture development - A review
The production of therapeutic proteins in mammalian cell culture is an essential unit operation in biopharmaceutical manufacture that can benefit from the predictive insights of effective process models, leading to accelerated process development and improved process control. This review outlines and evaluates current approaches to predictive model development for mammalian cell culture and protein production. Classical mechanistic and data driven approaches are analysed, together with potential challenges in model development and application, including the experimental requirements for parameter estimation. Hybrid models, which may offer greater robustness, are then explored along with hybrid model architecture and the steps involved in model development. Successful examples from other cell fermentation processes are also considered, for application to the development, monitoring and control of mammalian processes
Self-assembling nanomaterials: monitoring the formation of amyloid fibrils, with a focus on small-angle X-ray scattering.
Amyloid fibrils are attractive targets for applications in biotechnology. These thin, nanoscale protein fibers are highly ordered structures that self-assemble from their component proteins or peptides. This chapter describes the use of several biophysical techniques to monitor the formation of amyloid fibrils including a common dye-binding assay, turbidity assay, and small-angle X-ray scattering. These techniques provide information about the assembly mechanism, the rate and reproducibility of assembly, as well as the size of species along the assembly pathway
Functional fibrils derived from the peptide TTR1-cycloRGDfK that target cell adhesion and spreading
Marie N. Bongiovanni, Denis B. Scanlon, Sally L. Gra
Computational Modelling of Large Scale Phage Production Using a Two-Stage Batch Process
Cost effective and scalable methods for phage production are required to meet an increasing demand for phage, as an alternative to antibiotics. Computational models can assist the optimization of such production processes. A model is developed here that can simulate the dynamics of phage population growth and production in a two-stage, self-cycling process. The model incorporates variable infection parameters as a function of bacterial growth rate and employs ordinary differential equations, allowing application to a setup with multiple reactors. The model provides simple cost estimates as a function of key operational parameters including substrate concentration, feed volume and cycling times. For the phage and bacteria pairing examined, costs and productivity varied by three orders of magnitude, with the lowest cost found to be most sensitive to the influent substrate concentration and low level setting in the first vessel. An example case study of phage production is also presented, showing how parameter values affect the production costs and estimating production times. The approach presented is flexible and can be used to optimize phage production at laboratory or factory scale by minimizing costs or maximizing productivity
Exploiting amyloid: how and why bacteria use cross-β fibrils.
Many bacteria produce protein fibrils that are structurally analogous to those associated with protein misfolding diseases such as Alzheimer's disease. However, unlike fibrils associated with disease, bacterial amyloids have beneficial functions including conferring stability to biofilms, regulating development or imparting virulence. In the present review, we consider what makes amyloid fibrils so suitable for these roles and discuss recent developments in the study of bacterial amyloids, in particular the chaplins from Streptomyces coelicolor. We also consider the broader impact of the study of bacterial amyloids on our understanding of infection and disease and on developments in nanotechnology
Exploiting amyloid: how and why bacteria use cross-β fibrils. Biochemical Society Transactions. 2012; 40(4):728–734. [PubMed: 22817724
Abstract Many bacteria produce protein fibrils that are structurally analogous to those associated with protein misfolding diseases such as Alzheimer's disease. However, unlike fibrils associated with disease, bacterial amyloids have beneficial functions including conferring stability to biofilms, regulating development or imparting virulence. In the present review, we consider what makes amyloid fibrils so suitable for these roles and discuss recent developments in the study of bacterial amyloids, in particular the chaplins from Streptomyces coelicolor. We also consider the broader impact of the study of bacterial amyloids on our understanding of infection and disease and on developments in nanotechnology
- …