Uncovering the Oppenheimer Siddur: using scientific analysis to reveal the production process of a medieval illuminated Hebrew manuscript

The aim of this research was to use non-invasive scientifc analysis to uncover evidence of the planning process and relationship between pigments used in text copying and artwork production in the Oppenheimer Siddur (Oxford Bodleian Library MS Opp. 776), an illuminated 15th-century Hebrew prayer book. In many medieval Hebrew illuminated manuscripts, the authorship of the artwork is unknown. This manuscript’s colophon states that it was copied by its scribe-owner for personal family use but does not confrm who was responsible for the artwork. Prior deductive analysis suggested that the scribe-owner may also have been the manuscript’s artist, based on common motifs and an apparent shared colour palette appearing in both texts and artwork. Visual examination using high resolution digital images also identifed points of contact between pigments used in the manuscript’s texts and artwork, raising questions about the pigment application sequence, and concurrent versus sequential text copying and artwork production. An in-house developed remote spectral imaging system (PRISMS) with 10 flters spanning the spectral range from 400 to 880 nm was modifed for close-range application to image two of the folios to examine the sequence of production, identify the pigments and compare the materials used for the illumination and the text. Optical microscopy and Fourier Transform Infrared spectroscopy in the attenuated total refection mode (FTIR-ATR) were used directly on the folios to complement the spectral imaging data in binding media and pigment identifcation. The results revealed close matches in refectance spectra for the colorants and inks used in both text copying and illuminations, suggesting that the same mixture of colorants and inks have been used. The spectral imaging in the near infrared bands revealed a hidden underdrawing, indicating a design change during production of the manuscript, and the outlining of letters prior to coloured pigment being applied. The pigment use, the variation in the binder for diferent pigments and some elements of its production were found to be consistent with those described in historical sources. The evidence from this study supports the hypothesis that the scribe applied pigments for the manuscript’s artwork at the same time he did some of the scribal work which has implications for understandings of Jewish medieval visual cultures

Reversal of the ΔdegP Phenotypes by a Novel rpoE Allele of Escherichia coli

RseA sequesters RpoE (σE) to the inner membrane of Escherichia coli when envelope stress is low. Elevated envelope stress triggers RseA cleavage by the sequential action of two membrane proteases, DegS and RseP, releasing σE to activate an envelope stress reducing pathway. Revertants of a ΔdegP ΔbamB strain, which fails to grow at 37°C due to high envelope stress, harbored mutations in the rseA and rpoE genes. Null and missense rseA mutations constitutively hyper-activated the σE regulon and significantly reduced the major outer membrane protein (OMP) levels. In contrast, a novel rpoE allele, rpoE3, resulting from the partial duplication of the rpoE gene, increased σE levels greater than that seen in the rseA mutant background but did not reduce OMP levels. A σE-dependent RybB::LacZ construct showed only a weak activation of the σE pathway by rpoE3. Despite this, rpoE3 fully reversed the growth and envelope vesiculation phenotypes of ΔdegP. Interestingly, rpoE3 also brought down the modestly activated Cpx envelope stress pathway in the ΔdegP strain to the wild type level, showing the complementary nature of the σE and Cpx pathways. Through employing a labile mutant periplasmic protein, AcrAL222Q, it was determined that the rpoE3 mutation overcomes the ΔdegP phenotypes, in part, by activating a σE-dependent proteolytic pathway. Our data suggest that a reduction in the OMP levels is not intrinsic to the σE-mediated mechanism of lowering envelope stress. They also suggest that under extreme envelope stress, a tight homeostasis loop between RseA and σE may partly be responsible for cell death, and this loop can be broken by mutations that either lower RseA activity or increase σE levels

Periplasmic peptidyl-prolyl isomerases SurA and FkpA play an important role in the starvation-stress response (SSR) of Salmonella enterica serovar Typhimurium

Carbon-energy source (C)-starved cells of Salmonella enterica serovar Typhimurium (S. Typhimurium) are remarkably more resistant to stress than actively growing ones. Carbon-starved S. Typhimurium is capable of withstanding extended periods of starvation and assault from a number of different stresses that rapidly kill growing cells. These unique properties of the C-starved cell are the direct result of a series of genetic and physiological adaptations referred to as the starvation-stress response (SSR). Previous work established that the SSR of S. Typhimurium is partially regulated by the extracytoplasmic function sigma factor σE. As part of an effort to identify σE-regulated SSR genes, we investigated surA and fkpA, encoding two different classes of peptidyl-prolyl isomerase that function in folding cell envelope proteins. Both surA and fkpA are members of the heat-shock-inducible σE regulon of Escherichia coli. Although both genes are expressed in C-starved Salmonella cells, evidence indicates that surA and fkpA are not C-starvation-inducible. Furthermore, their expression during C-starvation does not appear to be σE-dependent. Nonetheless, surA and fkpA proved to be important, to differing degrees, for long-term C-starvation survival and for the cross-resistance of C-starved cells to high temperature, acidic pH, and the antimicrobial peptide polymyxin B, but neither were required for cross-resistance to oxidative stress. These results point to fundamental differences between heat-shock-inducible and C-starvation-inducible genes regulated by σE and suggest that genes other than surA and fkpA are involved in the σE-regulated branch of the SSR in Salmonella

Investigation of Yersinia pestis Laboratory Adaptation through a Combined Genomics and Proteomics Approach

The bacterial pathogen Yersinia pestis, the cause of plague in humans and animals, normally has a sylvatic lifestyle, cycling between fleas and mammals. In contrast, laboratory-grown Y. pestis experiences a more constant environment and conditions that it would not normally encounter. The transition from the natural environment to the laboratory results in a vastly different set of selective pressures, and represents what could be considered domestication. Understanding the kinds of adaptations Y. pestis undergoes as it becomes domesticated will contribute to understanding the basic biology of this important pathogen. In this study, we performed a parallel serial passage experiment (PSPE) to explore the mechanisms by which Y. pestis adapts to laboratory conditions, hypothesizing that cells would undergo significant changes in virulence and nutrient acquisition systems. Two wild strains were serially passaged in 12 independent populations each for ~750 generations, after which each population was analyzed using whole-genome sequencing, LC-MS/MS proteomic analysis, and GC/MS metabolomics. We observed considerable parallel evolution in the endpoint populations, detecting multiple independent mutations in ail, pepA, and zwf, suggesting that specific selective pressures are shaping evolutionary responses. Complementary LC-MS/MS proteomic data provide physiological context to the observed mutations, and reveal regulatory changes not necessarily associated with specific mutations, including changes in amino acid metabolism and cell envelope biogenesis. Proteomic data support hypotheses generated by genomic data in addition to suggesting future mechanistic studies, indicating that future whole-genome sequencing studies be designed to leverage proteomics as a critical complement