Structural diversity in the type IV pili of multidrug-resistant Acinetobacter

Acinetobacter baumannii is a Gram-negative coccobacillus found primarily in hospital settings that has recently emerged as a source of hospital-acquired infections. A. baumannii expresses a variety of virulence factors, including type IV pili, bacterial extracellular appendages often essential for attachment to host cells. Here, we report the high resolution structures of the major pilin subunit, PilA, from three Acinetobacter strains, demonstrating thatA. baumannii subsets produce morphologically distinct type IV pilin glycoproteins. We examine the consequences of this heterogeneity for protein folding and assembly as well as host-cell adhesion by Acinetobacter. Comparisons of genomic and structural data with pilin proteins from other species of soil gammaproteobacteria suggest that these structural differences stem from evolutionary pressure that has resulted in three distinct classes of type IVa pilins, each found in multiple species

The Rosetta All-Atom Energy Function for Macromolecular Modeling and Design

Over the past decade, the Rosetta biomolecular modeling suite has informed diverse biological questions and engineering challenges ranging from interpretation of low-resolution structural data to design of nanomaterials, protein therapeutics, and vaccines. Central to Rosetta's success is the energy function: a model parametrized from small-molecule and X-ray crystal structure data used to approximate the energy associated with each biomolecule conformation. This paper describes the mathematical models and physical concepts that underlie the latest Rosetta energy function, called the Rosetta Energy Function 2015 (REF15). Applying these concepts, we explain how to use Rosetta energies to identify and analyze the features of biomolecular models. Finally, we discuss the latest advances in the energy function that extend its capabilities from soluble proteins to also include membrane proteins, peptides containing noncanonical amino acids, small molecules, carbohydrates, nucleic acids, and other macromolecules

Better together: Elements of successful scientific software development in a distributed collaborative community

Many scientific disciplines rely on computational methods for data analysis, model generation, and prediction. Implementing these methods is often accomplished by researchers with domain expertise but without formal training in software engineering or computer science. This arrangement has led to underappreciation of sustainability and maintainability of scientific software tools developed in academic environments. Some software tools have avoided this fate, including the scientific library Rosetta. We use this software and its community as a case study to show how modern software development can be accomplished successfully, irrespective of subject area. Rosetta is one of the largest software suites for macromolecular modeling, with 3.1 million lines of code and many state-of-the-art applications. Since the mid 1990s, the software has been developed collaboratively by the RosettaCommons, a community of academics from over 60 institutions worldwide with diverse backgrounds including chemistry, biology, physiology, physics, engineering, mathematics, and computer science. Developing this software suite has provided us with more than two decades of experience in how to effectively develop advanced scientific software in a global community with hundreds of contributors. Here we illustrate the functioning of this development community by addressing technical aspects (like version control, testing, and maintenance), community-building strategies, diversity efforts, software dissemination, and user support. We demonstrate how modern computational research can thrive in a distributed collaborative community. The practices described here are independent of subject area and can be readily adopted by other software development communities

A Comprehensive, High-Resolution Map of a Gene’s Fitness Landscape

Mutations are central to evolution, providing the genetic variation upon which selection acts. A mutation’s effect on the suitability of a gene to perform a particular function (gene fitness) can be positive, negative, or neutral. Knowledge of the distribution of fitness effects (DFE) of mutations is fundamental for understanding evolutionary dynamics, molecular-level genetic variation, complex genetic disease, the accumulation of deleterious mutations, and the molecular clock. We present comprehensive DFEs for point and codon mutants of the Escherichia coli TEM-1 β-lactamase gene and missense mutations in the TEM-1 protein. These DFEs provide insight into the inherent benefits of the genetic code’s architecture, support for the hypothesis that mRNA stability dictates codon usage at the beginning of genes, an extensive framework for understanding protein mutational tolerance, and evidence that mutational effects on protein thermodynamic stability shape the DFE. Contrary to prevailing expectations, we find that deleterious effects of mutation primarily arise from a decrease in specific protein activity and not cellular protein levels

Characterization of a Fungal Thioesterase Having Claisen Cyclase and Deacetylase Activities in Melanin Biosynthesis

SummaryMelanins are a broad class of darkly pigmented macromolecules formed by oxidative polymerization of phenolic monomers. In fungi, melanins are known virulence factors that contribute to pathogenicity. Their biosynthesis generally involves polymerization of 1,8-dihydroxynaphthalene via a 1,3,6,8-tetrahydroxynaphthalene (THN) precursor assembled by multidomain, nonreducing polyketide synthases. Convergent routes to THN have evolved in fungi. Parallel heptaketide and hexaketide pathways exist that utilize conventional C-terminal thioesterase/Claisen cyclase domains and separate side-chain deacylases. Here, in vitro characterization of Pks1 from Colletotrichum lagenarium establishes a true THN synthase with a bifunctional thioesterase (TE) catalyzing both cyclization and deacetylation of an enzyme-bound hexaketide substrate. Chimeric TE domains were generated by swapping lid regions of active sites between classes of melanin TEs to gain insight into this unprecedented catalysis of carbon–carbon bond making and breaking by an α/β-hydrolase fold enzyme