Resistance to Mucosal Lysozyme Compensates for the Fitness Deficit of Peptidoglycan Modifications by Streptococcus pneumoniae

The abundance of lysozyme on mucosal surfaces suggests that successful colonizers must be able to evade its antimicrobial effects. Lysozyme has a muramidase activity that hydrolyzes bacterial peptidoglycan and a non-muramidase activity attributable to its function as a cationic antimicrobial peptide. Two enzymes (PgdA, a N-acetylglucosamine deacetylase, and Adr, an O-acetyl transferase) that modify different sites on the peptidoglycan of Streptococcus pneumoniae have been implicated in its resistance to lysozyme in vitro. Here we show that the antimicrobial effect of human lysozyme is due to its muramidase activity and that both peptidoglycan modifications are required for full resistance by pneumococci. To examine the contribution of lysozyme and peptidoglycan modifications during colonization of the upper respiratory tract, competition experiments were performed with wild-type and pgdAadr mutant pneumococci in lysozyme M-sufficient (LysM+/+) and -deficient (LysM−/−) mice. The wild-type strain out-competed the double mutant in LysM+/+, but not LysM−/− mice, indicating the importance of resistance to the muramidase activity of lysozyme during mucosal colonization. In contrast, strains containing single mutations in either pgdA or adr prevailed over the wild-type strain in both LysM+/+ and LysM−/− mice. Our findings demonstrate that individual peptidoglycan modifications diminish fitness during colonization. The competitive advantage of wild-type pneumococci in LysM+/+ but not LysM−/− mice suggests that the combination of peptidoglycan modifications reduces overall fitness, but that this is outweighed by the benefits of resistance to the peptidoglycan degrading activity of lysozyme

Isolation and crystallization of CP47, a Photosystem II chlorophyll binding protein. Degradation of CP47 upon dissociation from the core complex

The CP47 protein was isolated from Photosystem II membranes by using a combination of the detergents n-dodecyl-beta-D-maltoside and octyl-beta-D-thioglucoside. The purified CP47 was used in a series of crystallization experiments, which yielded highly reproducible hexagonal crystals. Immunoblot analysis revealed that the isolated CP47 undergoes degradation even under dim light conditions. This degradation takes place after the protein has been dissociated from the core complex. Proteolysis experiments with trypsin demonstrated that the dissociation of the CP47 from the PS II core complex results in changes that render the protein sensitive to proteolysis

Isolation and crystallization of CP47, a Photosystem II chlorophyll binding protein. Degradation of CP47 upon dissociation from the core complex

The CP47 protein was isolated from Photosystem II membranes by using a combination of the detergents n-dodecyl-beta-D-maltoside and octyl-beta-D-thioglucoside. The purified CP47 was used in a series of crystallization experiments, which yielded highly reproducible hexagonal crystals. Immunoblot analysis revealed that the isolated CP47 undergoes degradation even under dim light conditions. This degradation takes place after the protein has been dissociated from the core complex. Proteolysis experiments with trypsin demonstrated that the dissociation of the CP47 from the PS II core complex results in changes that render the protein sensitive to proteolysis. [References: 28

Association between dietary intake and nutritional status in Eastern Mediterranean patients receiving hemodialysis

BACKGROUND: Studies on hemodialysis (HD) patients reveal suboptimal dietary intakes, which have been linked to protein-energy wasting and its detrimental consequences. OBJECTIVE: Given the paucity of data regarding nutrient intakes of Eastern Mediterranean HD patients, we conducted a pilot study on HD patients at Heraklion, Crete, Greece to assess adequacy of dietary intakes and to determine their relationship with nutritional status. METHODS: Nutritional status of 36 patients aged 61.8±15.0 years was evaluated by three 24-hour dietary recalls, anthropometry and blood biochemical markers. RESULTS: The mean dietary energy and protein intakes were 34.4±2.1 kcal/kg and 1.25±0.067 g/kg, respectively. The anthropometric results were indicative of a well-maintained somatic status. 30.6% of the patients had adequate weight (BMI 18.5-24.9 kg/m2) and 69.4% were overweight or obese (BMI≥25 kg/m2). Patients had arm anthropometrics higher than the 25th percentile. Mean predialysis serum levels of urea and creatinine, were within the expected range, phosphorus was borderline high, while albumin and cholesterol were at the optimum level for HD patients. In univariate linear regression, a positive relation was observed between ideal weight-adjusted energy (wEI) and protein (wPI) intakes with anthropometric and biochemical indices. However, in the multivariate model, only the associations between dietary intakes of energy and protein with anthropometric indices remained significant. CONCLUSIONS: 24-hour derived-dietary intakes reached recommended targets and adequately reflected the nutritional status of the patients, according to anthropometric and biochemical indices. Additionally, the 24-hour recall method should be part of the routine care for HD patients, in order to identify patients at nutritional risk before objective parameters of wasting are documented. © 2020 - IOS Press and the authors. All rights reserved

Purification, crystallization and preliminary X-ray analysis of the peptidoglycan N-acetylglucosamine deacetylase BC1960 from Bacillus cereus in the presence of its substrate (GlcNAc)6

The peptidoglycan N-acetylglucosamine (GlcNAc) deacetylase BC1960 from Bacillus cereus (EC 3.5.1.33), an enzyme consisting of 275 amino acids, was crystallized in the presence of its substrate (GlcNAc)(6). The crystals belonged to the tetragonal space group P4(1)2(1)2, with unit-cell parameters a = b = 92.7, c = 242.9 A and four molecules in the asymmetric unit. A complete data set was collected at 100 K to a resolution of 2.38 A using synchrotron radiation

Purification, crystallization and preliminary X-ray analysis of the peptidoglycan N-acetylglucosamine deacetylase BC1960 from Bacillus cereus in the presence of its substrate (GlcNAc)6

The peptidoglycan N-acetylglucosamine (GlcNAc) deacetylase BC1960 from Bacillus cereus (EC 3.5.1.33), an enzyme consisting of 275 amino acids, was crystallized in the presence of its substrate (GlcNAc)(6). The crystals belonged to the tetragonal space group P4(1)2(1)2, with unit-cell parameters a = b = 92.7, c = 242.9 A and four molecules in the asymmetric unit. A complete data set was collected at 100 K to a resolution of 2.38 A using synchrotron radiation

Purification, crystallization and preliminary characterization of a putative LmbE-like deacetylase from Bacillus cereus

The BC1534 protein from B. cereus was purified and crystallized and a native X-ray diffraction data set was collected to 2.5 Å using synchrotron radiation