Proteomic evidence of dietary sources in ancient dental calculus.

Archaeological dental calculus has emerged as a rich source of ancient biomolecules, including proteins. Previous analyses of proteins extracted from ancient dental calculus revealed the presence of the dietary milk protein β-lactoglobulin, providing direct evidence of dairy consumption in the archaeological record. However, the potential for calculus to preserve other food-related proteins has not yet been systematically explored. Here we analyse shotgun metaproteomic data from 100 archaeological dental calculus samples ranging from the Iron Age to the post-medieval period (eighth century BC to nineteenth century AD) in England, as well as 14 dental calculus samples from contemporary dental patients and recently deceased individuals, to characterize the range and extent of dietary proteins preserved in dental calculus. In addition to milk proteins, we detect proteomic evidence of foodstuffs such as cereals and plant products, as well as the digestive enzyme salivary amylase. We discuss the importance of optimized protein extraction methods, data analysis approaches and authentication strategies in the identification of dietary proteins from archaeological dental calculus. This study demonstrates that proteomic approaches can robustly identify foodstuffs in the archaeological record that are typically under-represented due to their poor macroscopic preservation

The role of forensic anthropology in disaster victim identification (DVI):recent developments and future prospects

Forensic anthropological knowledge has been used in disaster victim identification (DVI) for over a century, but over the past decades, there have been a number of disaster events which have seen an increasing role for the forensic anthropologist. The experiences gained from some of the latest DVI operations have provided valuable lessons that have had an effect on the role and perceived value of the forensic anthropologist as part of the team managing the DVI process. This paper provides an overview of the ways in which forensic anthropologists may contribute to DVI with emphasis on how recent experiences and developments in forensic anthropology have augmented these contributions. Consequently, this paper reviews the value of forensic anthropological expertise at the disaster scene and in the mortuary, and discusses the way in which forensic anthropologists may use imaging in DVI efforts. Tissue-sampling strategies for DNA analysis, especially in the case of disasters with a large amount of fragmented remains, are also discussed. Additionally, consideration is given to the identification of survivors; the statistical basis of identification; the challenges related to some specific disaster scenarios; and education and training. Although forensic anthropologists can play a valuable role in different phases of a DVI operation, they never practice in isolation. The DVI process requires a multidisciplinary approach and, therefore, has a close collaboration with a range of forensic specialists

Improving catalytic function by ProSAR-driven enzyme evolution

We describe a directed evolution approach that should find broad application in generating enzymes that meet predefined process-design criteria. It augments recombination-based directed evolution by incorporating a strategy for statistical analysis of protein sequence activity relationships (ProSAR). This combination facilitates mutation-oriented enzyme optimization by permitting the capture of additional information contained in the sequence-activity data. The method thus enables identification of beneficial mutations even in variants with reduced function. We use this hybrid approach to evolve a bacterial halohydrin dehalogenase that improves the volumetric productivity of a cyanation process B4,000-fold. This improvement was required to meet the practical design criteria for a commercially relevant biocatalytic process involved in the synthesis of a cholesterollowering drug, atorvastatin (Lipitor), and was obtained by variants that had at least 35 mutations. Although interest in the use of enzymes as biocatalysts for chemical applications is increasing 1,2 , the performance of natural enzymes is rarely adequate for commercially viable processes. Many enzyme properties, such as specific activity, stability, chemo-and enantioselectivity, susceptibility to substrate and/or product inhibition and sensitivity to rapidly changing conditions, are difficult to optimize using rational design. Despite advances in protein engineering, there is a continuing need for more efficient and effective methods to improve multiple biochemical characteristics of an enzyme simultaneously 3,4 . Directed evolution is a powerful tool for protein optimization. The most efficient methods combine multiple rounds of diversity generation and gene recombination with functional screening to identify improved variants. The iterative nature of this approach results in stepwise improvements in overall function, yielding substantial improvements in desired enzyme properties Directed evolution can be performed in a variety of ways 10 , which are distinguished, for example, by the approach to generating libraries from available diversity. The field of in vitro recombination-based directed evolution 9 has focused on alternative methods for generating gene libraries rather than fundamentally altering the efficiency of the evolutionary process 7,10 , and despite advances, there remains a need to make the process of increasing enzyme function more efficient 3,4 . Our approach involves focusing the selective pressure on the mutations themselves rather than on the mutated gene. Quantitative structure-activity relationships (QSAR) have been used extensively in small-molecule 11 and peptide optimization 12 and have been proposed to be useful in protein engineering This study applies the concepts of QSAR to the problem of enzyme engineering. A multivariate protein optimization strategy based on protein sequence activity relationships (ProSAR) We projected that an economically viable process could be achieved if the following design criteria were met: complete conversion (100%) of at least 100 g per liter substrate, a volumetric productivity of 420 g product per liter per hour per gram of biocatalyst, a simple HN isolation procedure to recover high quality product in high yield, and a simple enzyme formulation process that obviates the need for extensive enzyme purification. We first expressed an Agrobacterium radiobacter HHDH in Escherichia coli and showed that it catalyzed the conversion of ethyl (S)-4-chloro-3-hydroxybutyrate (ECHB) to HN at neutral p

Insights into preventive measures for dental erosion

Dental erosion is defined as the loss of tooth substance by acid exposure not involving bacteria. The etiology of erosion is related to different behavioral, biological and chemical factors. Based on an overview of the current literature, this paper presents a summary of the preventive strategies relevant for patients suffering from dental erosion. Behavioral factors, such as special drinking habits, unhealthy lifestyle factors or occupational acid exposure, might modify the extent of dental erosion. Thus, preventive strategies have to include measures to reduce the frequency and duration of acid exposure as well as adequate oral hygiene measures, as it is known that eroded surfaces are more susceptible to abrasion. Biological factors, such as saliva or acquired pellicle, act protectively against erosive demineralization. Therefore, the production of saliva should be enhanced, especially in patients with hyposalivation or xerostomia. With regard to chemical factors, the modification of acidic solutions with ions, especially calcium, was shown to reduce the demineralization, but the efficacy depends on the other chemical factors, such as the type of acid. To enhance the remineralization of eroded surfaces and to prevent further progression of dental wear, high-concentrated fluoride applications are recommended. Currently, little information is available about the efficacy of other preventive strategies, such as calcium and laser application, as well as the use of matrix metalloproteinase inhibitors. Further studies considering these factors are required. In conclusion, preventive strategies for patients suffering from erosion are mainly obtained from in vitro and in situ studies and include dietary counseling, stimulation of salivary flow, optimization of fluoride regimens, modification of erosive beverages and adequate oral hygiene measures