Biology subjects

Beer, M.

Publications and source records attributed to Beer, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Systematic genome analysis enables long-overdue taxonomic reorganization of genus Orthoflavivirus; potential assignment of over 50 new species

Members of the Orthoflavivirus genus in the family Flaviviridae are primarily vector-transmitted RNA viruses infecting humans, other mammals and birds, including major pathogens such as tick borne encephalitis, West Nile, Zika and dengue viruses. Vector-borne flaviviruses are assigned to the Euflavivirus subgenus of orthoflaviviruses, with more divergent largely unclassified insect-specific flaviviruses, including cell fusing agent virus assigned to the second subgenus, Fusivirus. The currently classified 57 orthoflavivirus species have traditionally been assigned descriptively on the basis of their genome and antigenic relatedness, ecology and host and vector tropisms, although many more orthoflavi-like viruses remain unassigned. To investigate whether species might be demarcated using a fixed genetic divergence threshold, we examined distributions of pairwise distances in NS5 and E genes (which encode the RNA-dependent RNA polymerase and envelope protein, respectively) of classified orthoflaviviruses. Demarcation thresholds of 0.1 in the two regions would be most parsimonious with current euflavivirus classification. Exceptions include species in the tick-borne encephalitis virus complex with divergences below the threshold, and dengue virus types currently assigned to a single species that are substantially above. Application of the proposed thresholds would provide the means to establish 25 new euflavivirus and 33 new fusivirus species and reconcile several anomalies and inconsistencies in the current species taxonomy. The use of a species demarcation threshold brings consistency with the classification of viruses in other families. A robust and evolutionarily consistent framework furthermore enables taxonomic assignments of the numerous currently unassigned orthoflaviviruses that lack the phenotypic descriptions required for the current species definition.

microbiology↗

bioRxiv2026

Bat-borne influenza A H9N2 virus shows species-specific replication and transmission

Bat-borne H9N2 IAV (bat H9N2), initially detected in bat feces in Egypt in 2017, has recently attracted attention because of its unexpected pre-pandemic properties, including airborne transmission between ferrets and efficient replication in human lung explant cultures. However, its host range and potential reservoir hosts remain poorly defined, particularly with respect to additional bat hosts and mammalian or avian mixing-vessel species. Here, we compare the susceptibility of two pteropodid bat species, Rousettus aegyptiacus and Eidolon helvum, and additionally assessed pigs and quails following intranasal inoculation with bat H9N2. Juvenile R. aegyptiacus showed only low-level viral replication and shedding, largely restricted to the upper respiratory tract, and did not show transmission to contact bats. In contrast, both juvenile and adult E. helvum, despite showing no clinical signs, showed high-level viral replication and shedding in the upper respiratory tract, with efficient and early transmission to contact bats. Viral RNA was most abundant in the respiratory turbinates, the nasopharyngeal meatus and the tracheal epithelium, where it was associated with moderate to severe necrotizing rhinitis and tracheitis. The lower respiratory tract was less involved, with viral RNA confined to scattered bronchioles and associated with focal necrotizing bronchiolitis. Other organs, including the gastrointestinal tract, lacked histologic lesions and viral RNA labeling. In pigs and quails, intranasal inoculation did not result in detectable replication, transmission, or seroconversion. Our findings demonstrate marked differences in the susceptibility to bat H9N2 between the two fruit bat species tested and identify E. helvum as a highly susceptible host capable of rapid and efficient transmission. Conversely, pigs and quails were not susceptible, suggesting that additional viral adaptations may be required to allow for a productive infection in these species.

microbiology↗

bioRxiv2026

Two incursions, two viruses: Emergence of a second novel Shamonda virus clade in Central Europe, 2026

Following the emergence of Shamonda virus (SHAV) in Europe in 2026, we identified a second, genetically distinct SHAV clade in cattle in Germany. Consistent differences across all three genome segments together with divergent regional distribution patterns indicate independent introductions. We have designated the two clades as SHAV Europe 1 (SHAV-EU1) and SHAV Europe 2 (SHAV-EU2). The unexpected co-circulation of these two clades has important implications for diagnostics, surveillance, host range assessment, risk evaluation, and control measures.

microbiology↗

bioRxiv2026

Experimental reproduction numbers disentangle vaccine effects on susceptibility and infectiousness during H5N1 transmission in geese

Vaccination against high pathogenicity avian influenza virus (HPAIV) is increasingly used to protect poultry, but vaccine performance is commonly inferred from clinical protection and virus shedding rather than measured transmission. We asked whether reproduction numbers from controlled transmission experiments can quantify how vaccination changes susceptibility and infectiousness. Domestic geese were prime-boost vaccinated with an H5 clade 2.3.4.4b RNA-replicon vaccine and challenged with homologous HPAIV H5N1. Replicated seeder-sentinel groups represented transmission among unvaccinated animals, to vaccinated contacts, and from vaccinated breakthrough-infected animals. Vaccinated directly challenged geese remained clinically protected although all became RT-qPCR-positive. Estimated reproduction numbers were R0=4.7 (95% CI, 2.8-8.0) among unvaccinated geese, Rs=2.6 (1.6-4.5) for transmission to vaccinated contacts, Ri=3.7 (2.0-6.8) for transmission from vaccinated infected geese, and Rvacc=2.0 (0.8-4.9) for a fully vaccinated population. Vaccination reduced transmission but did not reduce the point estimate for Rvacc below one under these intensive exposure conditions. Vaccinated infected geese also shed substantially less viral RNA, whereas the estimated reduction in infectiousness was more modest, indicating that RNA shedding alone may not reliably predict transmission reduction. Experimental reproduction numbers therefore provide a direct population-level complement to conventional vaccine endpoints and separate effects on susceptibility from effects on onward transmission.

microbiology↗

bioRxiv2026

A transition state-like acylenzyme conformation distinguishes carbapenemase activity in class A β-lactamases

Carbapenems are the most potent {beta}-lactams, key antibiotics for healthcare-associated infections by Gram-negative bacteria and evade hydrolysis by most {beta}-lactamases, but are increasingly threatened by emergence of enzymes exhibiting hydrolytic activity towards them. Of the four recognised {beta}-lactamase subclasses, class A (active-site serine enzymes that hydrolyse {beta}-lactams via a covalent acylenzyme intermediate) is the most widely disseminated and, while the majority of such enzymes react with carbapenems to form long-lasting acylenzyme complexes, several possess carbapenem-hydrolyzing activity (carbapenemases). Here, we investigate the basis for these differences in a panel of class A {beta}-lactamases using molecular dynamics (MD) simulations of the respective acylenzyme complexes and tetrahedral intermediates (TI). The simulations reveal multiple features associated with catalytic activity across the spectrum of enzymes tested, including more extensive interactions of the carbapenem acylenzyme carbonyl and generally increased lifetimes of active site water molecules positioned for deacylation. Analysis of the dynamic trajectories shows carbapenemases to have reduced root mean-squared fluctuation (RMSF) differences between the acylenzyme and TI, that are not limited to the active site, indicating that the acylenzyme complex is pre-organised for reaction in carbapenemases but not in carbapenem-inhibited enzymes. Similarly, Principal Component Analysis (PCA) of acylenzyme and TI dynamics shows greater overlap between the two states in carbapenemases, providing further evidence for acylenzyme pre-organisation. Such simulations may represent an effective computational assay able to identify enzymes with carbapenemase activity at relatively modest computational cost.

biochemistry↗

bioRxiv2026

It happened again - emergence of a Shamonda-like orthobunyavirus in Central Europe, 2026

Since mid-June 2026, an acute, non-fatal syndrome with high herd morbidity, reduced milk yield, diarrhea, fever and lethargy affected dairy cattle in southern Germany and Switzerland. Major viral pathogens were excluded. Pan-Simbuvirus PCR first detected an orthobunyavirus, subsequently confirmed by metagenomic sequencing, which enabled recovery of a complete "Shamonda-like" genome. One sentence summary lineMore than a decade after the emergence of Schmallenberg virus in 2011, a novel "Shamonda-like" orthobunyavirus of the Simbu serogroup has now emerged in Central Europe.

microbiology↗

bioRxiv2026

A scalable mesh microelectrode array platform for longitudinal electrophysiology in neural spheroids

Electrophysiological interfacing remains difficult in three-dimensional in vitro models, when using planar microelectrode arrays or optical methods. This challenge limits experimental progress using such models, despite their promise of better physiological relevance than monolayer cell culture. Mesh MEAs which can integrate conformally on or even within tissue offer a possible solution but are not yet widely accessible. Here, we present a mesh MEA device, designed as a simple, manufacturable platform for spheroid electrophysiology. In neural spheroids, the device enabled longitudinal electrophysiological recordings and pharmacological modulation of spontaneous electrical activity. On native polyimide meshes, spheroids maintained their shape while cells enveloped the mesh, embedding electrodes to a depth of 100 {micro}m after 2 weeks. In contrast, laminin biofunctionalization of the mesh promoted outgrowth and migration of cells. This device and associated methods should be adaptable to organoids, ex vivo tissue, or bioengineered in vitro models.

neuroscience↗

bioRxiv2026

First evaluation of a human DPP4 transgenic hamster model for MERS-CoV pathogenesis and transmission

MERS-CoV poses a constant pandemic risk, as its viral lineages continue evolving, and zoonotic spillover events could lead to random viral polymorphisms that might lead to human adapted variants. Currently, no small animal model reliably recapitulates both disease progression and transmission dynamics, which are critical aspects for counter-viral measures like vaccine development. Although the Syrian hamster is an optimal animal model for SARS-CoV-2 infection and transmission, it is naturally resistant to MERS-CoV infection. Dipeptidyl peptidase-4 (DPP4) is the functional receptor for MERS-CoV infection, and is highly expressed in human kidney, intestine, liver, and lung tissues. Here, we evaluated the suitability of a human DPP4 (hDPP4) transgenic Syrian hamster model for MERS-CoV research. We used two different MERS-CoV strains (EMC/2012 and D10540/2023) for intranasal inoculation of hamsters. Both strains replicated efficiently, led to comparable severe clinical outcomes, and had similar viral transmission efficiencies. MERS-CoV RNA and nucleoprotein antigen were mainly detected in the brain and the respiratory tract. In summary, we validated a novel hDPP4-transgenic hamster as a suitable model for MERS-CoV infection enabling vaccine and transmission research.

microbiology↗

bioRxiv2026

Acyl-enzyme dynamics, tautomerisation and hydration regulate turnover of carbapenem antibiotics by the OXA-48 β-lactamase

OXA-48 is a globally disseminated class D serine {beta}-lactamase that efficiently confers resistance to a range of {beta}-lactam antibiotics, including carbapenems, the most potent such agents versus Enterobacterales (Escherichia coli and relatives). Here we characterise the interactions of OXA-48 with the acyl-enzyme complex intermediates formed on its reaction with the carbapenems meropenem and ertapenem using X-ray crystallography and molecular dynamics (MD) simulations. X-ray crystal structures identify acyl-enzymes in both the {Delta}1-imine and {Delta}2-enamine pyrroline tautomeric forms. MD simulations show the epimeric {Delta}2 tautomers of meropenem and ertapenem to more frequently adopt binding poses competent for hydrolysis, i.e. with an appropriate orientation of the carbapenem 6-hydroxyethyl group and positioning of the water molecule required for deacylation; the results indicate that the {Delta}2 tautomers are preferred for deacylation over the {Delta}1-tautomer. MD simulations based on the crystal structures show that, compared to OXA-48, acyl-enzyme complexes of OXA-519 (a natural OXA-48 variant with a single Val120Leu substitution adjacent to the catalytic general base) more frequently sampled conformations favouring hydrolysis, or formation of the alternative {beta}-lactone deacylation product. MD simulations of complexes derived from quantum mechanics/molecular mechanics (QM/MM) simulations show the meropenem-derived {beta}-lactone product is better retained in the OXA-48 active site than hydrolysed meropenem, consistent with reversible {beta}-lactone formation. Overall, our results demonstrate how acyl-enzyme tautomerisation, dynamics and hydration collectively modulate degradation of 1{beta}-methyl carbapenems by class D {beta}-lactamases of the OXA-48 group, and how subtle changes in active site structure potentiate such effects in the OXA-519 variant.

biochemistry↗

bioRxiv2026

Buffaloed in Brandenburg: Germany's first Brush with Foot-and-Mouth Disease after four Decades of Freedom

Foot-and-mouth disease (FMD) virus is one of the most feared and most consequential pathogens of livestock worldwide. It can be spread rapidly by the transboundary movement of animals, animal products and byproducts. In January 2025, Germany detected its first FMD outbreak since 1988 in extensively reared water buffalo on a small farm in the state of Brandenburg, directly outside Berlin, the federal capital. Immediate control measures including a standstill for movements of susceptible animals and pre-emptive culling were implemented by the veterinary authorities. Whole-genome sequencing identified the virus as serotype O, topotype ME-SA, lineage SA-2018 and revealed extensive recombination, but cross-neutralization assays suggested good heterologous protection by an O/PanAsia-2 vaccine strain. Epidemiological back-calculation placed the time of virus introduction in late December 2024. Although the entry route remains unresolved, human-associated introduction is most likely. Network analysis revealed minimal farm connectivity, and simulations predicted low potential for onward transmission, which is consistent with the outbreak being ultimately restricted to a single herd. This event underscores the constant and unpredictable risk of introduction of the virus. Early detection through increased awareness and comprehensive differential diagnostics as well as the international collaboration of veterinary services, laboratories and experts are essential in the face of the global presence of FMD.

microbiology↗

bioRxiv2026

Dispersal, adaptation and persistence of H5N1 in the sub-Antarctic and Antarctica

High pathogenicity avian influenza virus (HPAIV) H5N1 reached the sub-Antarctic and Antarctica in 2023, subsequently spreading to remote locations within this region where it had devastating impacts on seal, penguin and albatross populations. The threat to marine wildlife over this broad area exemplifies the need to understand H5N1 long-distance dispersal and evolution. We obtained 104 novel viral genomic sequences from samples that we collected at South Georgia, Kerguelen, Crozet, Prince Edward, Falklands/Malvinas Islands and the Antarctic Peninsula in a region spanning 8,000 kilometers. Using recent phylogeographic modeling advances we show that H5N1 spread encompassed numerous transmission events between distant locations, accumulating mammalian-adaptive mutations in the process. Seals are the most affected species, but we reveal that the long-distance eastward virus dispersal better aligns with the long-distance movements of large petrels and albatrosses. The risk of H5N1 endemisation, dispersal to other locations and ongoing evolution are highly concerning.

microbiology↗

bioRxiv2026

One and Done: A safe, adaptable single-cycle SARS-CoV-2 vaccine platform blocks XBB.1.5 infection and transmission

A next-generation SARS-CoV-2 vaccine must address the currently inadequate prevention of virus transmission, particularly against emerging variants of concern, a challenge that none of the licensed commercial vaccines fully meet. Effective control of respiratory pandemics necessitates vaccines that 1) can be rapidly adapted, 2) have high patient compliance with simple and non-invasive administration, and 3) block transmission in a virus challenge. We describe here the characterization of an updated single-cycle SARS-CoV-2 vaccine candidate (scVac), engineered as a replication-defective virus with targeted deletions of the E gene and ORF6 and ORF7a, along with a truncation of ORF3a. The candidate carries an Omicron XBB.1.5 Spike (scVacXBB), maintaining all essential antigenic properties. The vaccine demonstrated an excellent safety profile in K18-hACE2 transgenic mice, the most sensitive virulence model, with no clinical signs or adverse events observed. In the Syrian hamster model, potent systemic and mucosal immune responses were induced, along with a strong neutralizing antibody response. Notably, there was no virus transmission to co-housed naive animals, which outperforms a bivalent Omicron mRNA vaccine reference. Our results demonstrate that scVacXBB-induced immunity not only prevents disease but also effectively blocks transmission. Furthermore, the successful introduction of the XBB.1.5 Spike protein into the scVac platform demonstrates the pipelines ability to adapt quickly to any emerging variant. These findings highlight the potential of this single-cycle concept as a next-generation COVID-19 vaccine, offering robust protection with a strong safety profile.

immunology↗

bioRxiv2026

Development of a multi-species luciferase-based double antigen ELISA for the detection of antibodies against Influenza A virus H5 clade 2.3.4.4b

The highly pathogenic avian influenza viruses (HPAIV) of subtype H5N1 represent a major threat to animal and public health. The current panzootic with H5 clade 2.3.4.4b has caused numerous, widespread outbreaks in various domestic and wild avian species with high mortalities, massive losses and a high frequency of spillover events to unexpected novel mammalian hosts such as dairy cows. The global H5N1 situation raises serious concerns about zoonotic risks due to effective mammal-to-mammal transmission. Therefore, it is critical to increase surveillance intensity of a broadened species range, particularly at the human-animal interface. For this purpose, reliable and cost-effective serological tools that are easy to perform and suitable for high-throughput screenings are critically needed. The newly developed double antigen ELISA format employing a luminescence-based detection technology has demonstrated to comply with such prerequisites. The assay allowed the detection of H5-specific antibodies even early after infection or vaccination in a wide range of birds and mammals including humans. It further demonstrated superior analytical sensitivity and high specificity for antibodies directed against H5 hemagglutinin of clade 2.3.4.4b as no cross-reactivity with other hemagglutinin subtypes was observed. Thus, the assay represents a valuable contribution to existing serological diagnostic tests for a clade-optimized detection of influenza A virus antibodies in a broad range of species. ImportanceThe ongoing HPAIV H5N1 panzootic has caused numerous outbreaks in domestic and wild animals with frequent spillover events to unexpected host species, which underscores the importance of an intensified surveillance. However, sensitive and specific multi-species serological assays represent a major gap. For this purpose, we developed a novel double antigen ELISA which employs an innovative luminescence-based read-out strategy. The test allowed a highly sensitive and specific detection of H5-specific antibodies even early after infection or vaccination in a wide range of avian and mammalian species including humans. It therefore represents a significant contribution to improving species-independent serological diagnostic tools for the detection of influenza A virus antibodies.

microbiology↗

bioRxiv2025

Time-resolved ligand dynamics revealed in a β-lactamase using room-temperature serial crystallography

{beta}-Lactamases catalyze {beta}-lactam antibiotic hydrolysis and are important contributors to bacterial antimicrobial resistance; {beta}-lactamase inhibitors are widely used to overcome {beta}-lactamase-mediated antibiotic resistance. Nucleophilic serine {beta}-lactamases (SBLs) react with their substrates and clinically available inhibitors via a covalent reaction to give complexes which can undergo further reaction. Using room temperature drop on fixed target serial crystallography, where ligands are rapidly mixed with microcrystals, and classical single-crystal crystallography at cryogenic temperatures, we investigate the reversible covalent reaction of the SBL CTX-M-15 with the diazobicyclooctane inhibitor avibactam. We observe avibactam covalently reacted (ring-opened) with the nucleophilic Ser70, at timepoints from 80 ms to minutes (room temperature) and hours (100 K). These crystallographic data reveal time-dependent movement of the avibactam carbamoyl complex, from 1.3 s onwards, that has implications for the 5-exo-trig recyclization mechanism that determines inhibitor reformation. Combined with molecular dynamics simulations and quantum mechanics calculations at the density functional theory level, the results show that in the first seconds of the reaction the avibactam N-sulfate nitrogen is poorly positioned for recyclization. This subsequently equilibrates after 10 s to a stable endpoint that is in a conformation potentially primed to initiate recyclization through attack of the N-sulfate nitrogen on the carbamoyl carbon. These results further demonstrate the capacity of room-temperature serial crystallography to capture time-resolved changes in ligand conformation at an enzyme active site, complementing discrete classical cryo-crystallography. These data inform on ligand dynamics and the stereoelectronics of diazobicyclooctane inhibition, aiding drug discovery efforts to develop inhibitors of nucleophilic enzymes.

biochemistry↗

bioRxiv2025

Drop on fixed target reaction initiation approach for serial and time resolved crystallography

We describe the design and implementation of a drop on fixed target method for time-resolved serial crystallography at both synchrotron and XFEL facilities. A piezoelectric droplet dispensing pipette is employed for addition of picolitre volume (40 - 90 pL) aqueous droplets, containing (co-)substrate(s) or ligand(s), onto enzyme microcrystals immobilised on a solid support. The system was tested with various enzyme systems, including lysozyme and two -lactamases, CTX-M-15 and AmpCEC. Mitigation strategies for cross-well contamination, including the implementation of interleaved controls, are described; the overall performance of the system at synchrotron and X-ray free electron laser facilities was evaluated. This drop on fixed target method is a reliable framework for time-resolved crystallography and will improve the consistency of measurements across facilities.

biophysics↗

bioRxiv2025

Mass mortality at penguin mega-colonies due to avian cholera confounds H5N1 HPAIV surveillance in Antarctica

In the austral summer 2023/2024, H5N1 high pathogenicity avian influenza virus (HPAIV) was reported for the first time in Antarctica. Concerns of HPAIV causing high mortality of seabirds and mammals prompted immediate efforts to track its spread and impact on endemic wildlife. In March 2024, we visited the Danger Islands archipelago, that hosts two mega-colonies of Adelie penguins, and observed an unusual mortality estimated in thousands of Adelie penguins and other species. Swabs and tissues were collected for molecular detection of infectious agents from 49 carcasses, and additional tissues for histology from a selection of 9 carcasses. We unexpectedly detected Pasteurella multocida DNA in 46 of 49 individuals, and diagnosed avian cholera, and not HPAI, as the cause of death of most of these animals. By metagenomics, we retrieved the genomic sequences of the Pasteurella multocida strain which caused the epizootic, and the phylogenetic analysis showed a close relation with strains previously reported in the Southern Ocean area. This study confirms avian cholera as a relevant cause of mortality in the Antarctic region, and overall highlights the importance of considering avian cholera in the differential diagnoses during mortality events in Antarctica, even with the concurrent circulation of HPAIV.

microbiology↗

bioRxiv2025

High pathogenicity avian influenza virus H5N1 (clade 2.3.4.4b) drives mass mortality in Eurasian crane (Grus grus) populations in Germany, 2025

In autumn 2025, an unprecedented mass mortality event was observed among the western migrating subpopulation of Eurasian cranes (Grus grus) in Germany. Systemic infection with highly pathogenic avian influenza virus H5N1, clade 2.3.4.4b, genotype DI.2.1, was identified as the cause of acute death. The gregarious behaviour of cranes at feeding and resting sites likely has contributed to the rapid and massive dissemination of viruses within the crane population.

ecology↗

bioRxiv2025

A dynamical-nonequilibrium molecular dynamics (D-NEMD) alanine scanning approach for identifying allosteric positions in proteins

Allosteric effects are widespread in proteins, but predicting the impact of sequence substitutions at positions distant from ligand-binding or enzyme active sites remains challenging, as their effects are mediated through complex dynamical networks. Pinpointing such positions experimentally is labour-intensive and low throughput. Equilibrium molecular dynamics (MD) simulations are useful: analysis methods can identify functional distal sites and networks, while free energy simulations can predict effects on stability or binding, given sufficient sampling; such simulations are typically time-consuming, requiring extensive simulation of each individual mutant. Here, we introduce a dynamical-nonequilibrium MD (D-NEMD) protocol using alanine substitutions as targeted perturbations to identify distal positions that influence enzyme function. Using the well characterised class A {beta}-lactamase KPC-2 as a test system, we show that D-NEMD distinguishes functional from non-functional sites based on whether substitutions generate structured, long-range responses that reach the active site. KPC-2 is clinically important due to its broad substrate spectrum and the emergence of resistance-associated point mutations, many of which act through non-local effects on catalytic residues. Alanine substitutions at positions 179 and 164, which disrupt the {Omega}-loop salt bridge, trigger persistent structural responses propagating through the protein and reach both catalytic residues and the oxyanion-hole backbone. Likewise, alanine substitution at position 220 elicits pronounced responses extending to the active-site {beta}-sheet and key loops, consistent with its known role in substrate specificity. In contrast, substitution at position 276--mutations of which have negligible kinetic impact--produces only local displacements with minimal propagation and no effect on catalytically relevant regions. These differential response patterns align with experimentally observed resistance phenotypes. D-NEMD therefore provides a fast, generalisable, and predictive approach for identifying allosterically connected distal positions. The protocol complements equilibrium MD, is straightforward to implement, and offers a tractable route for prioritising candidate sites for mechanistic study or future mutational scanning efforts. Statement of significanceWe introduce a dynamical-nonequilibrium MD (D-NEMD) protocol that uses alanine substitutions as the perturbation to identify residues, particularly those distal to an active or binding site, that modulate biological activity. We test this approach on the clinically important {beta}-lactamase KPC-2: it distinguishes positions that elicit long-range responses that reach catalytic residues from another with only local effects, in agreement with experiment. The D-NEMD approach allows for tests of statistical robustness of responses to mutation, enabling prioritisation of candidate residues for experimental mutation. This should aid in identifying distal sites that may influence enzyme activity or serve other allosteric roles. The approach is generalisable to other proteins of biomedical and biotechnological interest.

biochemistry↗