Obtaining well-ordered crystals is a major hurdle to X-ray structure determination of membrane proteins. To facilitate crystal optimization, we investigated the detergent stability of 24 eukaryotic and prokaryotic membrane proteins, predominantly transporters, using a fluorescent-based unfolding assay. We have benchmarked the stability required for crystallization in small micelle detergents, as they are statistically more likely to lead to high-resolution structures. Using this information, we have been able to obtain well-diffracting crystals for a number of sodium and proton-dependent transporters. By including in the analysis seven membrane proteins for which structures are already known, AmtB, GlpG, Mhp1, GlpT, EmrD, NhaA, and LacY, it was further possible to demonstrate an overall trend between protein stability and structural resolution. We suggest that by monitoring membrane protein stability with reference to the benchmarks described here, greater efforts can be placed on constructs and conditions more likely to yield high-resolution structures.

Antibiotic resistance is a global health crisis that requires urgent action to stop its spread. To counteract the spread of antibiotic resistance, we must improve our understanding of the origin and spread of resistant bacteria in both community and healthcare settings. Unfortunately, little attention is being given to contain the spread of antibiotic resistance in community settings (i.e., locations outside of a hospital inpatient, acute care setting, or a hospital clinic setting), despite some studies have consistently reported a high prevalence of antibiotic resistance in the community settings. This study aimed to investigate the prevalence of antibiotic resistance in commensal Escherichia coli isolates from healthy humans in community settings in LMICs. Using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, we synthesized studies conducted from 1989 to May 2020. A total of 9363 articles were obtained from the search and prevalence data were extracted from 33 articles and pooled together. This gave a pooled prevalence of antibiotic resistance (top ten antibiotics commonly prescribed in LMICs) in commensal E. coli isolates from human sources in community settings in LMICs of: ampicillin (72% of 13,531 isolates, 95% CI: 65–79), cefotaxime (27% of 6700 isolates, 95% CI: 12–44), chloramphenicol (45% of 7012 isolates, 95% CI: 35–53), ciprofloxacin (17% of 10,618 isolates, 95% CI: 11–25), co-trimoxazole (63% of 10,561 isolates, 95% CI: 52–73), nalidixic acid (30% of 9819 isolates, 95% CI: 21–40), oxytetracycline (78% of 1451 isolates, 95% CI: 65–88), streptomycin (58% of 3831 isolates, 95% CI: 44–72), tetracycline (67% of 11,847 isolates, 95% CI: 59–74), and trimethoprim (67% of 3265 isolates, 95% CI: 59–75). Here, we provided an appraisal of the evidence of the high prevalence of antibiotic resistance by commensal E. coli in community settings in LMICs. Our findings will have important ramifications for public health policy design to contain the spread of antibiotic resistance in community settings. Indeed, commensal E. coli is the main reservoir for spreading antibiotic resistance to other pathogenic enteric bacteria via mobile genetic elements.

Membrane bilayers are made up of a myriad of different lipids that regulate the functional activity, stability, and oligomerization of many membrane proteins. Despite their importance, screening the structural and functional impact of lipid–protein interactions to identify specific lipid requirements remains a major challenge. Here, we use the FSEC-TS assay to show cardiolipin-dependent stabilization of the dimeric sodium/proton antiporter NhaA, demonstrating its ability to detect specific protein-lipid interactions. Based on the principle of FSEC-TS, we then engineer a simple thermal-shift assay (GFP-TS), which facilitates the high-throughput screening of lipid- and ligand- interactions with membrane proteins. By comparing the thermostability of medically relevant eukaryotic membrane proteins and a selection of bacterial counterparts, we reveal that eukaryotic proteins appear to have evolved to be more dependent to the presence of specific lipids.