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319‐44 is a human monoclonal antibody capable of passively protecting mice against tick‐mediated infection with Borreliella burgdorferi, the bacterial genospecies responsible for Lyme disease in North America. In vitro, 319‐44 has complement‐dependent borreliacidal activity and spirochete agglutinating properties. Here, we report the 2.2 Å‐resolution crystal structure of 319‐44 Fab fragments in complex with Outer surface protein A (OspA), the ~30 kDa lipoprotein that was the basis of the first‐generation Lyme disease vaccine approved in the United States. The 319‐44 epitope is focused on OspA β‐strands 19, 20, and 21, and the loops between β‐strands 16‐17, 18‐19, and 20‐21. Contact with loop 20‐21 explains competition with LA‐2, the murine monoclonal antibody used to estimate serum borreliacidal activities in the first‐generation Lyme disease vaccine clinical trials. A high‐resolution B‐cell epitope map of OspA will accelerate structure‐based design of second generation OspA‐based vaccines. [ABSTRACT FROM AUTHOR]

In this report we investigated, within a group of closely related single domain camelid antibodies (VHHs), the relationship between binding affinity and neutralizing activity as it pertains to ricin, a fast-acting toxin and biothreat agent. The V1C7-like VHHs (V1C7, V2B9, V2E8, and V5C1) are similar in amino acid sequence, but differ in their binding affinities and toxin-neutralizing activities. Using the X-ray crystal structure of V1C7 in complex with ricin's enzymatic subunit (RTA) as a template, Rosetta-based homology modeling coupled with energetic decomposition led us to predict that a single pairwise interaction between Arg29 on V5C1 and Glu67 on RTA was responsible for the difference in ricin toxin binding affinity between V1C7, a weak neutralizer, and V5C1, a moderate neutralizer. This prediction was borne out experimentally: substitution of Arg for Gly at position 29 enhanced V1C7's binding affinity for ricin, whereas the reverse (ie, Gly for Arg at position 29) diminished V5C1's binding affinity by >10 fold. As expected, the V5C1R29G mutant was largely devoid of toxin-neutralizing activity (TNA). However, the TNA of the V1C7G29R mutant was not correspondingly improved, indicating that in the V1C7 family binding affinity alone does not account for differences in antibody function. V1C7 and V5C1, as well as their respective point mutants, recognized indistinguishable epitopes on RTA, at least at the level of sensitivity afforded by hydrogen-deuterium mass spectrometry. The results of this study have implications for engineering therapeutic antibodies because they demonstrate that even subtle differences in epitope specificity can account for important differences in antibody function. [ABSTRACT FROM AUTHOR]

Irreversible inhibition of the essential nervous system enzyme acetylcholinesterase by organophosphate nerve agents and pesticides may quickly lead to death. Oxime reactivators currently used as antidotes are generally less effective against pesticide exposure than nerve agent exposure, and pesticide exposure constitutes the majority of cases of organophosphate poisoning in the world. The current lack of published structural data specific to human acetylcholinesterase organophosphateinhibited and oxime-bound states hinders development of effective medical treatments. We have solved structures of human acetylcholinesterase in different states in complex with the organophosphate insecticide, paraoxon, and oximes. Reaction with paraoxon results in a highly perturbed acyl loop that causes a narrowing of the gorge in the peripheral site that may impede entry of reactivators. This appears characteristic of acetylcholinesterase inhibition by organophosphate insecticides but not nerve agents. Additional changes seen at the dimer interface are novel and provide further examples of the disruptive effect of paraoxon. Ternary structures of paraoxon-inhibited human acetylcholinesterase in complex with the oximes HI6 and 2-PAM reveals relatively poor positioning for reactivation. This study provides a structural foundation for improved reactivator design for the treatment of organophosphate intoxication. [ABSTRACT FROM AUTHOR]

ABSTRACT In this report, we describe the X-ray crystal structures of two single domain camelid antibodies (VHH), F5 and F8, each in complex with ricin toxin's enzymatic subunit (RTA). F5 has potent toxin-neutralizing activity, while F8 has weak neutralizing activity. F5 buried a total of 1760 Å2 in complex with RTA and made contact with three prominent secondary structural elements: α-helix B (Residues 98-106), β-strand h (Residues 113-117), and the C-terminus of α-helix D (Residues 154-156). F8 buried 1103 Å2 in complex with RTA that was centered primarily on β-strand h. As such, the structural epitope of F8 is essentially nested within that of F5. All three of the F5 complementarity determining regions CDRs were involved in RTA contact, whereas F8 interactions were almost entirely mediated by CDR3, which essentially formed a seventh β-strand within RTA's centrally located β-sheet. A comparison of the two structures reported here to several previously reported (RTA-VHH) structures identifies putative contact sites on RTA, particularly α-helix B, associated with potent toxin-neutralizing activity. This information has implications for rational design of RTA-based subunit vaccines for biodefense. Proteins 2016; 84:1162-1172. © 2016 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Coxiella burnetii is a highly infectious bacterium and potential agent of bioterrorism. However, it has not been studied as extensively as other biological agents, and very few of its proteins have been structurally characterized. To address this situation, we undertook a study of critical metabolic enzymes in C. burnetii that have great potential as drug targets. We used high-throughput techniques to produce novel crystal structures of 48 of these proteins. We selected one protein, C. burnetii dihydrofolate reductase (CbDHFR), for additional work to demonstrate the value of these structures for structure-based drug design. This enzyme's structure reveals a feature in the substrate binding groove that is different between CbDHFR and human dihydrofolate reductase (hDHFR). We then identified a compound by in silico screening that exploits this binding groove difference, and demonstrated that this compound inhibits CbDHFR with at least 25-fold greater potency than hDHFR. Since this binding groove feature is shared by many other prokaryotes, the compound identified could form the basis of a novel antibacterial agent effective against a broad spectrum of pathogenic bacteria. [ABSTRACT FROM AUTHOR]

Service-learning has become an important component in the education of medical and dental students around the world. Dental students at the University of Witwatersrand, South Africa, provide dental services to rural communities via the Phelophepa train or a Public Oral Health Facility (POHF). The Phelophepa train is a mobile primary health care facility offering dental, pharmacy, nursing, and medical services provided by health professions students. The objective of this study was to determine the impact this experience with service-learning has had on dental students. Final-year dental students in 2008 and 2009 participated in the study by completing a self-administered questionnaire. There was a 100 percent response rate (N=55) on the demographic questions and a 98 percent response rate (N=54) on the opinion questions. Students on the Phelophepa train performed more extractions and examinations than those at the POHFs. Most students (95 percent), both on the train and at the clinics, reported that their clinical skills and efficiency had improved, and 96 percent felt more aware of the communities' needs. Almost all the students (96 percent) reported that the experience had helped them define their personal strengths and weaknesses. Complaints they mentioned included large numbers of patients (87 percent), long working hours (60 percent), and equipment being inadequate (4S percent) or not working (40 percent). This program positively impacted these students and enhanced their personal growth and social responsibility by exposing them to the needs of rural and urban communities in South Africa. [ABSTRACT FROM AUTHOR]

AMP-activated protein kinase (AMPK) is a master metabolic regulator for controlling cellular energy homeostasis. Its homolog in yeast, SNF1, is activated in response to glucose depletion and other stresses. The catalytic (α) subunit of AMPK/SNF1 in yeast (Snf1) contains a protein Ser/Thr kinase domain (KD), an auto-inhibitory domain (AID) and a region that mediates interactions with the two regulatory (β and γ) subunits. Here, the crystal structure of residues 41-440 of Snf1, which include the KD and AID, is reported at 2.4 Å resolution. The AID is completely disordered in the crystal. A new inhibited conformation of the KD is observed in a DFG-out conformation and with the glycine-rich loop adopting a structure that blocks ATP binding to the active site. [ABSTRACT FROM AUTHOR]

Compulsory community service (CCS) was introduced into the health service by the South African government to address the shortage and maldistribution of health professionals within the public sector. The aim of this study was to determine the attitudes of dental therapy students regarding CCS. A self-administered questionnaire was delivered to the two dental schools that train dental therapists in South Africa. There was a 64 percent response rate; 56 percent of the respondents were female. The aver- age age was 20.3 years. There was no difference in the variables between the two dental schools, so the results were combined. The majority (81 percent) supported the introduction of CCS and preferred to carry it out in Kwa-Zulu Natal. Most students opted to perform oral health promotional (64 percent) and clinical (15 percent) activities. By aligning these requirements with the current dental needs and priority strategies of the South African Department of Health, this support would add much value to the delivery of oral health services. [ABSTRACT FROM AUTHOR]

Investigates the crystal structure of the oxidized human sulfite oxidase cytochrome b[sub 5] domain. Electron transport between the molybdenum cofactor of sulfite oxidase and the heme of cytochrome c; Crystallography; Distinct structural features present in the domain.

The severe personnel shortage in the health professions in most developing communities is well documented. This dearth of health workers and the widespread adoption of the primary health care approach (PHCA), requires that health professionals be trained to understand and effectively utilize the skills offered by other disciplines in the health care field. Nurses are expected to play active roles in the promotion of health including oral health, particularly in the rural under-served communities. There is a paucity of oral health education in nursing curricula. This article describes a 4-day module on oral health promotion developed and delivered to 36 nurses as part of a 12-month primary health care nursing (PHCN) training course. The module utilized a variety of educational methods and materials to facilitate and encourage both individual and group learning. The module was evaluated using both student academic attainment and perceptions as outcome measures. The evaluation and the experience of facilitating this module show that an oral health promotion module of short duration can sensitize nurses to the importance of oral health and increase oral health knowledge and skills. [ABSTRACT FROM AUTHOR]

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The molybdenum- and iron-containing enzyme sulfite oxidase catalyzes the physiologically vital oxidation of sulfite to sulfate. Sulfite oxidase contains three domains: an N-terminal cytochrome b(5) domain, a central domain harboring the molybdenum cofactor (Moco) and a C-terminal dimerization domain. Oxidation of the substrate sulfite is coupled to the transfer of two electrons to the molybdenum cofactor. Subsequently, these electrons are passed on, one at a time, to the b(5) heme of sulfite oxidase and from there to the soluble electron carrier cytochrome c. The crystal structure of the oxidized human sulfite oxidase cytochrome b(5) domain has been determined at 1.2 A resolution and has been refined to a crystallographic R factor of 0.107 (R(free) = 0.137). A comparison of this structure with other b(5)-type cytochromes reveals distinct structural features present in the sulfite oxidase b(5) domain which promote optimal electron transport between the Moco of sulfite oxidase and the heme of cytochrome c.