Serum albumin (SA) is the most abundant plasma protein in mammals. SA is a multifunctional protein with extraordinary ligand binding capacity, making it a transporter molecule for a diverse range of metabolites, drugs, nutrients, metals and other molecules. Due to its ligand binding properties, albumins have wide clinical, pharmaceutical, and biochemical applications. Albumins are also allergenic, and exhibit a high degree of cross-reactivity due to significant sequence and structure similarity of SAs from different organisms. Here we present crystal structures of albumins from cattle (BSA), horse (ESA) and rabbit (RSA) serums. The structural data are correlated with the results of immunological studies of SAs. We also analyze the conservation or divergence of structures and sequences of SAs in the context of their potential allergenicity and cross-reactivity. In addition, we identified a previously uncharacterized ligand binding site in the structure of RSA, and calcium binding sites in the structure of BSA, which is the first serum albumin structure to contain metal ions.

Metals are essential in many biological processes, and metal ions are modeled in roughly 40% of the macromolecular structures in the Protein Data Bank (PDB). However, a significant fraction of these structures contain poorly modeled metal-binding sites. CheckMyMetal (CMM) is an easy-to-use metal-binding site validation server for macromolecules that is freely available at http://csgid.org/ csgid/metal_sites. The CMM server can detect incorrect metal assignments as well as geometrical and other irregularities in the metal-binding sites. Guidelines for metal-site modeling and validation in macromolecules are illustrated by several practical examples grouped by the type of metal. These examples show CMM users (and crystallographers in general) problems they may encounter during the modeling of a specific metal ion.

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The low reproducibility of published experimental results in many scientific disciplines has recently garnered negative attention in scientific journals and the general media. Public transparency, including the availability of `raw' experimental data, will help to address growing concerns regarding scientific integrity. Macromolecular X-ray crystallography has led the way in requiring the public dissemination of atomic coordinates and a wealth of experimental data, making the field one of the most reproducible in the biological sciences. However, there remains no mandate for public disclosure of the original diffraction data. The Integrated Resource for Reproducibility in Macromolecular Crystallography (IRRMC) has been developed to archive raw data from diffraction experiments and, equally importantly, to provide related metadata. Currently, the database of our resource contains data from 2920 macromolecular diffraction experiments (5767 data sets), accounting for around 3% of all depositions in the Protein Data Bank (PDB), with their corresponding partially curated metadata. IRRMC utilizes distributed storage implemented using a federated architecture of many independent storage servers, which provides both scalability and sustainability. The resource, which is accessibleviathe web portal at http://www.proteindiffraction.org, can be searched using various criteria. All data are available for unrestricted access and download. The resource serves as a proof of concept and demonstrates the feasibility of archiving raw diffraction data and associated metadata from X-ray crystallographic studies of biological macromolecules. The goal is to expand this resource and include data sets that failed to yield X-ray structures in order to facilitate collaborative efforts that will improve protein structure-determination methods and to ensure the availability of `orphan' data left behind for various reasons by individual investigators and/or extinct structural genomics projects.

Background Alternaria is one of the most common molds associated with allergic diseases and 80% of Alternaria-sensitive patients produce IgE antibodies to a major protein allergen, Alt a 1. The structure and function of Alt a 1 is unknown. Objective To obtain a high resolution structure of Alt a 1 by X-ray crystallography and to investigate structural relationships between Alt a 1 and other allergens and proteins reported in the Protein Data Bank. Methods X-ray crystallography was used to determine the structure of Alt a 1 using a custom-designed set of crystallization conditions. An initial Alt a 1 model was determined by the application of a Ta6Br122+ cluster and Single-wavelength Anomalous Diffraction. Bioinformatic analyses were used to compare the Alt a 1 sequence and structure with other proteins. Results Alt a 1 is a unique β-barrel comprising 11 β-strands and forms a ‘butterfly-like’ dimer linked by a single disulfide bond, with a large (1345Å2) dimer interface. Intramolecular disulfide bonds are conserved among Alt a 1 homologs. Currently, the Alt a 1 structure has no equivalent in the Protein Data Bank. Bioinformatics analyses suggest that the structure is found exclusively in fungi. Four previously reported putative IgE binding peptides have been located on the Alt a 1 structure. Conclusions Alt a 1 has a unique, dimeric β-barrel structure that appears to define a new protein family with unknown function found exclusively in fungi. The location of IgE antibody binding epitopes is in agreement with the structural analysis of Alt a 1.The Alt a 1 structure will allow mechanistic structure/function studies and immunologic studies directed towards new forms of immunotherapy for Alternaria-sensitive allergic patients.

Background Albumins are multifunctional proteins present in the blood serum of animals. They can bind and transport a wide variety of ligands which they accommodate due to their conformational flexibility. Serum albumins are highly conserved both in amino acid sequence and three-dimensional structure. Several mammalian and avian serum albumins (SAs) are also allergens. Sensitization to one of the SAs coupled with the high degree of conservation between SAs may result in cross-reactive antibodies in allergic individuals. Sensitivity to SA generally begins with exposure to an aeroallergen, which can then lead to cross-sensitization to serum albumins present in food. Scope of Review This review focuses on the allergenicity of SAs presented in a structural context. Major Conclusions SA allergenicity is unusual taking into account the high sequence identity and similarity between SA from different species and human serum albumin. Cross-reactivity of human antibodies towards different SAs is one of the most important characteristics of these allergens. General Significance Establishing a relationship between sequence and structure of different SAs and their interactions with antibodies is crucial for understanding the mechanisms of cross-sensitization of atopic individuals. Structural information can also lead to better design and production of recombinant SAs to replace natural proteins in allergy testing and desensitization. Therefore, structural analyses are important for diagnostic and treatment purposes.

Survivin, a subunit of the chromosome passenger complex (CPC), binds the N-terminal tail of histone H3, which is phosphorylated on T3 by Haspin kinase, and localizes the complex to the inner centromeres. We used x-ray crystallography to determine the residues of Survivin that are important in binding phosphomodified histone H3. Mutation of amino acids that interact with the histone N-terminus lowered in vitro tail binding affinity and reduced CPC recruitment to the inner centromere in cells, validating our solved structures. Phylogenetic analysis shows that nonmammalian vertebrates have two Survivin paralogues, which we name class A and B. A distinguishing feature of these paralogues is an H-to-R change in an amino acid that interacts with the histone T3 phosphate. The binding to histone tails of the human class A paralogue, which has a histidine at this position, is sensitive to changes around physiological pH, whereas Xenopus Survivin class B is less so. Our data demonstrate that Survivin paralogues have different characteristics of phosphospecific binding to threonine-3 of histone H3, providing new insight into the biology of the inner centromere.

The massive technical and computational progress of biomolecular crystallography has generated some adverse side effects. Most crystal structure models, produced by crystallographers or well‐trained structural biologists, constitute useful sources of information, but occasional extreme outliers remind us that the process of structure determination is not fail‐safe. The occurrence of severe errors or gross misinterpretations raises fundamental questions: Why do such aberrations emerge in the first place? How did they evade the sophisticated validation procedures which often produce clear and dire warnings, and why were severe errors not noticed by the depositors themselves, their supervisors, referees and editors? Once detected, what can be done to either correct, improve or eliminate such models? How do incorrect models affect the underlying claims or biomedical hypotheses they were intended, but failed, to support? What is the long‐range effect of the propagation of such errors? And finally, what mechanisms can be envisioned to restore the validity of the scientific record and, if necessary, retract publications that are clearly invalidated by the lack of experimental evidence? We suggest that cognitive bias and flawed epistemology are likely at the root of the problem. By using examples from the published literature and from public repositories such as the Protein Data Bank, we provide case summaries to guide correction or improvement of structural models. When strong claims are unsustainable because of a deficient crystallographic model, removal of such a model and even retraction of the affected publication are necessary to restore the integrity of the scientific record.