Macrolide-specific efflux pump MacAB-TolC has been identified in diverse Gram-negative bacteria including Escherichia coli. The inner membrane transporter MacB requires the outer membrane factor TolC and the periplasmic adaptor protein MacA to form a functional tripartite complex. In this study, we used a chimeric protein containing the tip region of the TolC ␣-barrel to investigate the role of the TolC ␣-barrel tip region with regard to its interaction with MacA. The chimeric protein formed a stable complex with MacA, and the complex formation was abolished by substitution at the functionally essential residues located at the MacA ␣-helical tip region. Electron microscopic study delineated that this complex was made by tip-to-tip interaction between the tip regions of the ␣-barrels of TolC and MacA, which correlated well with the TolC and MacA complex calculated by molecular dynamics. Taken together, our results demonstrate that the MacA hexamer interacts with TolC in a tip-to-tip manner, and implies the manner by which MacA induces opening of the TolC channel.Drug resistance of microbial pathogens presents an increasing threat to public health (1). In Gram-negative pathogens, high levels of intrinsic or acquired drug resistance are conferred by three-component multidrug efflux pumps, which are composed of the inner membrane transporter, the outer membrane factor (OMF), and the periplasmic membrane fusion protein (MFP) 4 (2-5). These tripartite complexes span the entire twomembrane envelope of Gram-negative bacteria and expel various molecules into the medium, utilizing a proton gradient or ATP hydrolysis. The inner membrane transporters belong to one of three structurally dissimilar superfamilies of proteins: resistance-nodulation-cell division (RND), ATP-binding cassette (ABC), or major facilitator. The inner membrane transporters expel the substrates through the central channel of the OMF, as exemplified by Escherichia coli TolC, which spans the outer membrane (6). The MFP, which connects the other two components in the periplasm, is also essential for the function of the efflux pump.In E. coli, AcrAB-TolC acts as a major multidrug efflux pump (7-9), where AcrB is the RND-type inner membrane transporter and AcrA belongs to MFP. The homotrimeric TolC is embedded in the outer membrane and continues ϳ100 Å into the periplasmic space as an ␣-barrel composed of six ␣-hairpins that form the wall of a 35-Å inner-diameter cylindrical channel (10). The TolC channel is closed at the aperture end and the channel opening is induced only in the presence of the other components, the mechanism of which remains to be determined at the molecular level.The MacAB-TolC pump has been identified in E. coli; the inner membrane transporter MacB belongs to non-canonic ABC-type transporters (8,9,11,12), and MFP MacA shares structural similarity with AcrA (sequence similarity 44%) (13). Overproduction of MacAB results in increased resistance to the macrolide antibiotics in macrolide-susceptible AcrAB-deficient E. coli (8, 9, 11).The s...

Introduction The clinical characteristics of skin were investigated to study the inter‐relationship and changes in the biophysical properties of the epidermal and dermal layers associated with aging using noninvasive methods. Methods Our study included 100 healthy women aged between the early 20s and late 60s. Biophysical characteristics of skin such as color (brightness and spots), transparency, wrinkle on crow's feet, elasticity, hydration, sebum content, glossiness, and transepidermal water loss measured under controlled conditions. Results This study performed in a Korean population demonstrated that aging significantly affects human skin in terms of parameters such as wrinkles, skin color, elasticity, and epidermal hydration. Age‐related changes in skin hydration showed varying patterns between the epidermis and dermis. Skin color showed heterogeneous characteristics between the upper and lower epidermal layers associated with aging. Skin elasticity and wrinkles were observed to show and inversely proportional relationship in the early 40s. Conclusions We confirmed the significant influence of aging on the biophysical properties of skin and determined the distinct age‐related biophysical changes in the epidermal and dermal layers of skin using noninvasive method. This study indicates the need for further research to investigate the distinctive age‐related changes in characteristics of the epidermal and dermal layers of human skin.

Lysozymes are the first line of defense for a diverse range of organisms that catalyze the degradation of bacterial peptidoglycan. Gram-negative bacteria produce proteinaceous lysozyme inhibitors to protect themselves from the action of lysozymes. To date, MliC or PliC (membrane-bound or periplasmic inhibitor of c-type lysozyme, respectively) has been found in various Gram-negative bacteria. Here, we report the crystal structures of Brucella abortus PliC and its complex with human c-type lysozyme. The complex structure demonstrates that the invariant loop of MliC/PliC plays a crucial role in the inhibition of lysozyme via its insertion into the active site cleft of the lysozyme, as previously observed in the complex structure of Pseudomonas aeruginosa MliC and chicken c-type lysozyme. We identified a new binding interface between a loop adjacent to the active site of human lysozyme and a loop carrying Glu112 of B. abortus PliC, the structure of which was disordered in P. aeruginosa MliC. Because MliC/PliC family members have been implicated as putative colonization or virulence factors, the structures and mechanism of action of MliC/PliC will be relevant to the control of bacterial growth in animal hosts.

Background Striae distensae are common dermal lesions that progress through two different stages: the striae rubra, which appears to be erythematous, and striae alba, which is characterized by a hypopigmented feature. The clinical characteristics between striae distensae stages and normal skin remain unknown. Objectives We aimed to investigate the clinical characteristics according to stages of striae distensae in terms of their biophysical properties, using objective noninvasive measurements in comparison with adjacent normal skin. Methods Sixty‐one healthy female subjects with striae distensae were included as follows: 30 with striae rubra and 31 with striae alba on the abdomen and thighs. Hydration of the epidermis and dermis, skin color brightness, and Erythema index were measured. Skin elasticity, roughness, and dermal echo‐density of the skin with striae distensae and adjacent normal skin were also measured. Results Hydration of the epidermis and dermis showed no significant difference between the skin with striae distensae and normal skin. Brightness of skin with striae alba and normal skin was significantly higher than that of skin with striae rubra. Erythema index of skin with striae rubra was significantly higher than that of skin with striae alba and normal skin. Skin with striae rubra and striae alba had a rougher surface than normal skin. Elasticity and dermal echo‐density were significantly lower in striae distensae skin. Conclusions Striae rubra and striae alba had similar biophysical properties in terms of skin hydration, elasticity, roughness, and dermal density. Moreover, striae distensae have less elasticity, more roughness, and lower dermal density than normal skin.

CueP was initially identified as a copper-resistance gene in Salmonella enterica serovar Typhimurium, which has evolved to survive in the phagosomes of macrophages. Recently, CueP was determined to be a periplasmic copper-binding protein and has been implicated in the transfer of copper ions to SodCII in the periplasm. In this study, the crystal structure of CueP has been determined, revealing a V-shaped dimeric structure. The conserved cysteine and histidine residues are clustered on the surface of one side of the C-terminal domain, suggesting that this cysteine- and histidine-rich region is related to the function of CueP. LC-MS/MS analysis established the presence of a disulfide bond between Cys96 and Cys176 under aerobic conditions. Subsequent biophysical analyses showed that the CueP protein binds copper and zinc, and the mutation of Cys104 to serine (C104S) dramatically reduced the binding affinity for copper and zinc, suggesting that the cysteine- and histidine-rich cluster is responsible for copper binding. This study provides a structural basis for the participation of CueP in the resistance of the intracellular pathogen Salmonella to copper.