RESUMEN
BACKGROUND: Signaling proteins such as protein kinases adopt a diverse array of conformations to respond to regulatory signals in signaling pathways. Perhaps the most fundamental conformational change of a kinase is the transition between active and inactive states, and defining the conformational features associated with kinase activation is critical for selectively targeting abnormally regulated kinases in diseases. While manual examination of crystal structures have led to the identification of key structural features associated with kinase activation, the large number of kinase crystal structures (~3,500) and extensive conformational diversity displayed by the protein kinase superfamily poses unique challenges in fully defining the conformational features associated with kinase activation. Although some computational approaches have been proposed, they are typically based on a small subset of crystal structures using measurements biased towards the active site geometry. RESULTS: We utilize an unbiased informatics based machine learning approach to classify all eukaryotic protein kinase conformations deposited in the PDB. We show that the orientation of the activation segment, measured by φ, ψ, χ1, and pseudo-dihedral angles more accurately classify kinase crystal conformations than existing methods. We show that the formation of the K-E salt bridge is statistically dependent upon the activation segment orientation and identify evolutionary differences between the activation segment conformation of tyrosine and serine/threonine kinases. We provide evidence that our method can identify conformational changes associated with the binding of allosteric regulatory proteins, and show that the greatest variation in inactive structures comes from kinase group and family specific side chain orientations. CONCLUSION: We have provided the first comprehensive machine learning based classification of protein kinase active/inactive conformations, taking into account more structures and measurements than any previous classification effort. Further, our unbiased classification of inactive structures reveals residues associated with kinase functional specificity. To enable classification of new crystal structures, we have made our classifier publicly accessible through a stand-alone program housed at https://github.com/esbg/kinconform [DOI: 10.5281/zenodo.249090 ].
Asunto(s)
Aprendizaje Automático , Proteínas Quinasas/química , Dominio Catalítico , Modelos Moleculares , Conformación ProteicaRESUMEN
Protein kinases represent a large and diverse family of evolutionarily related proteins that are abnormally regulated in human cancers. Although genome sequencing studies have revealed thousands of variants in protein kinases, translating "big" genomic data into biological knowledge remains a challenge. Here, we describe an ontological framework for integrating and conceptualizing diverse forms of information related to kinase activation and regulatory mechanisms in a machine readable, human understandable form. We demonstrate the utility of this framework in analyzing the cancer kinome, and in generating testable hypotheses for experimental studies. Through the iterative process of aggregate ontology querying, hypothesis generation and experimental validation, we identify a novel mutational hotspot in the αC-ß4 loop of the kinase domain and demonstrate the functional impact of the identified variants in epidermal growth factor receptor (EGFR) constitutive activity and inhibitor sensitivity. We provide a unified resource for the kinase and cancer community, ProKinO, housed at http://vulcan.cs.uga.edu/prokino.
Asunto(s)
Neoplasias/enzimología , Proteínas Quinasas/genética , Secuencia de Aminoácidos , Animales , Antineoplásicos/farmacología , Células CHO , Dominio Catalítico , Cricetinae , Cricetulus , Minería de Datos , Gefitinib , Ontología de Genes , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Bases del Conocimiento , Modelos Moleculares , Neoplasias/genética , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas/química , Quinazolinas/farmacología , Alineación de Secuencia , Programas InformáticosRESUMEN
Dynamic interactions between gut microbiota and a host's innate and adaptive immune systems play key roles in maintaining intestinal homeostasis and inhibiting inflammation. The gut microbiota metabolizes proteins and complex carbohydrates, synthesize vitamins, and produce an enormous number of metabolic products that can mediate cross-talk between gut epithelial and immune cells. As a defense mechanism, gut epithelial cells produce a mucosal barrier to segregate microbiota from host immune cells and reduce intestinal permeability. An impaired interaction between gut microbiota and the mucosal immune system can lead to an increased abundance of potentially pathogenic gram-negative bacteria and their associated metabolic changes, disrupting the epithelial barrier and increasing susceptibility to infections. Gut dysbiosis, or negative alterations in gut microbial composition, can also dysregulate immune responses, causing inflammation, oxidative stress, and insulin resistance. Over time, chronic dysbiosis and the translocation of bacteria and their metabolic products across the mucosal barrier may increase prevalence of type 2 diabetes, cardiovascular disease, inflammatory bowel disease, autoimmune disease, and a variety of cancers. In this paper, we highlight the pivotal role gut microbiota and their metabolites (short-chain fatty acids (SCFAs)) play in mucosal immunity.
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Corrections have been made to "Gut Microbiota and Immune System Interactions" [...].
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Multiple sequence alignments (MSAs) are a fundamental analysis tool used throughout biology to investigate relationships between protein sequence, structure, function, evolutionary history, and patterns of disease-associated variants. However, their widespread application in systems biology research is currently hindered by the lack of user-friendly tools to simultaneously visualize, manipulate and query the information conceptualized in large sequence alignments, and the challenges in integrating MSAs with multiple orthogonal data such as cancer variants and post-translational modifications, which are often stored in heterogeneous data sources and formats. Here, we present the Multiple Sequence Alignment Ontology (MSAOnt), which represents a profile or consensus alignment in an ontological format. Subsets of the alignment are easily selected through the SPARQL Protocol and RDF Query Language for downstream statistical analysis or visualization. We have also created the Kinome Viewer (KinView), an interactive integrative visualization that places eukaryotic protein kinase cancer variants in the context of natural sequence variation and experimentally determined post-translational modifications, which play central roles in the regulation of cellular signaling pathways. Using KinView, we identified differential phosphorylation patterns between tyrosine and serine/threonine kinases in the activation segment, a major kinase regulatory region that is often mutated in proliferative diseases. We discuss cancer variants that disrupt phosphorylation sites in the activation segment, and show how KinView can be used as a comparative tool to identify differences and similarities in natural variation, cancer variants and post-translational modifications between kinase groups, families and subfamilies. Based on KinView comparisons, we identify and experimentally characterize a regulatory tyrosine (Y177PLK4) in the PLK4 C-terminal activation segment region termed the P+1 loop. To further demonstrate the application of KinView in hypothesis generation and testing, we formulate and validate a hypothesis explaining a novel predicted loss-of-function variant (D523NPKCß) in the regulatory spine of PKCß, a recently identified tumor suppressor kinase. KinView provides a novel, extensible interface for performing comparative analyses between subsets of kinases and for integrating multiple types of residue specific annotations in user friendly formats.