Recognition Pliability Is Coupled to Structural Heterogeneity: A Calmodulin Intrinsically Disordered Binding Region Complex

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Misincorporation of the proline homologue Aze (azetidine-2-carboxylic acid) into recombinant myelin basic protein.

peer reviewedWe have evaluated the effects of the proline homologue Aze (1) (azetidine-2-carboxylic acid) on growth of Escherichia coli strains used to over-express recombinant forms of murine myelin basic protein (rmMBP), and on the degree of misincorporation. Addition of Aze to minimal media resulted in severe diminution of growth rate, but rmMBP could still be produced and purified. Mass spectrometry indicated that a detectable proportion of the rmMBP produced had incorporated Aze instead of proline (Pro), to a maximum of three of eleven possible sites. Molecular modelling of a proline-rich region of rmMBP illustrated that the misincorporation of Aze at any site would cause a severe bend in the polypeptide chain, and that multiple Pro-->Aze substitutions would completely disrupt a poly-proline type II structure that has been conjectured to be functionally significant

The proline-rich region of 18.5 kDa myelin basic protein binds to the SH3-domain of Fyn tyrosine kinase with the aid of an upstream segment to form a dynamic complex in vitro

The intrinsically disordered 18.5 kDa classic isoform of MBP (myelin basic protein) interacts with Fyn kinase during oligodendrocyte development and myelination. It does so primarily via a central proline-rich SH3 (Src homology 3) ligand (T92–R104, murine 18.5 kDa MBP sequence numbering) that is part of a molecular switch due to its high degree of conservation and modification by MAP (mitogen-activated protein) and other kinases, especially at residues T92 and T95. Here, we show using co-transfection experiments of an early developmental oligodendroglial cell line (N19) that an MBP segment upstream of the primary ligand is involved in MBP–Fyn–SH3 association in cellula. Using solution NMR spectroscopy in vitro, we define this segment to comprise MBP residues (T62–L68), and demonstrate further that residues (V83–P93) are the predominant SH3-target, assessed by the degree of chemical shift change upon titration. We show by chemical shift index analysis that there is no formation of local poly-proline type II structure in the proline-rich segment upon binding, and by NOE (nuclear Overhauser effect) and relaxation measurements that MBP remains dynamic even while complexed with Fyn–SH3. The association is a new example first of a non-canonical SH3-domain interaction and second of a fuzzy MBP complex

Lyme Disease Frontiers: Reconciling Borrelia Biology and Clinical Conundrums

Lyme disease is a complex tick-borne zoonosis that poses an escalating public health threat in several parts of the world, despite sophisticated healthcare infrastructure and decades of effort to address the problem. Concepts like the true burden of the illness, from incidence rates to longstanding consequences of infection, and optimal case management, also remain shrouded in controversy. At the heart of this multidisciplinary issue are the causative spirochetal pathogens belonging to the Borrelia Lyme complex. Their unusual physiology and versatile lifestyle have challenged microbiologists, and may also hold the key to unlocking mysteries of the disease. The goal of this review is therefore to integrate established and emerging concepts of Borrelia biology and pathogenesis, and position them in the broader context of biomedical research and clinical practice. We begin by considering the conventions around diagnosing and characterizing Lyme disease that have served as a conceptual framework for the discipline. We then explore virulence from the perspective of both host (genetic and environmental predispositions) and pathogen (serotypes, dissemination, and immune modulation), as well as considering antimicrobial strategies (lab methodology, resistance, persistence, and clinical application), and borrelial adaptations of hypothesized medical significance (phenotypic plasticity or pleomorphy)

Profiling disease burden and Borrelia seroprevalence in Canadians with complex and chronic illness.

Lyme disease, caused by vector-borne Borrelia bacteria, can present with diverse multi-system symptoms that resemble other conditions. The objective of this study was to evaluate disease presentations and Borrelia seroreactivity in individuals experiencing a spectrum of chronic and complex illnesses. We recruited 157 participants from Eastern Canada who reported one or more diagnoses of Lyme disease, neurological, rheumatic, autoimmune, inflammatory, gastrointestinal, or cardiovascular illnesses, or were asymptomatic and presumed healthy. Intake categories were used to classify participants based on their perceived proximity to Lyme disease, distinguishing between those with a disclosed history of Borrelia infection, those with lookalike conditions (e.g. fibromyalgia syndrome), and those with unrelated ailments (e.g. intestinal polyps). Participants completed three questionnaires, the SF-36 v1, SIQR, and HMQ, to capture symptoms and functional burden, and provided blood serum for analysis at an accredited diagnostic lab. Two-tiered IgG and IgM serological assessments (whole cell ELISA and Western blot) were performed in a blinded fashion on all samples. The pattern of symptoms and functional burden were similarly profound in the presumptive Lyme and Lyme-like disease categories. Borrelia seroprevalence across the study cohort was 10% for each of IgG and IgM, and occurred within and beyond the Lyme disease intake category. Western blot positivity in the absence of reactive ELISA was also substantial. Fibromyalgia was the most common individual diagnostic tag disclosed by two-tier IgG-positive participants who did not report a history of Lyme disease. Within the IgG seropositive cohort, the presence of antibodies against the 31 kDa Outer Surface Protein A (OspA) was associated with significantly better health outcomes. Previously, this marker has been linked to treatment-refractory Lyme arthritis. Overall, our findings support prior observations of phenotypic overlap between Lyme and other diseases. Seropositivity associated with non-specific symptoms and functional impairment warrants further mechanistic investigation and therapeutic optimization