Structure, dynamics, and molecular inhibition of the Staphylococcus aureus m1A22-tRNA methyltransferase TrmK.

The enzyme m1A22-tRNA methyltransferase (TrmK) catalyzes the transfer of a methyl group to the N1 of adenine 22 in bacterial tRNAs. TrmK is essential for Staphylococcus aureus survival during infection but has no homolog in mammals, making it a promising target for antibiotic development. Here, we characterize the structure and function of S. aureus TrmK (SaTrmK) using X-ray crystallography, binding assays, and molecular dynamics simulations. We report crystal structures for the SaTrmK apoenzyme as well as in complexes with methyl donor SAM and co-product product SAH. Isothermal titration calorimetry showed that SAM binds to the enzyme with favorable but modest enthalpic and entropic contributions, whereas SAH binding leads to an entropic penalty compensated for by a large favorable enthalpic contribution. Molecular dynamics simulations point to specific motions of the C-terminal domain being altered by SAM binding, which might have implications for tRNA recruitment. In addition, activity assays for SaTrmK-catalyzed methylation of A22 mutants of tRNALeu demonstrate that the adenine at position 22 is absolutely essential. In silico screening of compounds suggested the multifunctional organic toxin plumbagin as a potential inhibitor of TrmK, which was confirmed by activity measurements. Furthermore, LC-MS data indicated the protein was covalently modified by one equivalent of the inhibitor, and proteolytic digestion coupled with LC-MS identified Cys92 in the vicinity of the SAM-binding site as the sole residue modified. These results identify a cryptic binding pocket of SaTrmK, laying a foundation for future structure-based drug discovery.

The Rural PILL Program: A Postdischarge Telepharmacy Intervention for Rural Veterans.

PURPOSE: To evaluate the efficacy of the Rural Pharmacological Intervention in Late Life (PILL) program, a quality improvement initiative in which a Boston-based pharmacist provided postdischarge telepharmacy care to veterans from rural Maine. METHODS: Using an automated screening tool, we identified 100 veterans aged 65 and older who had an acute care admission to VA medical centers in Boston or Maine and were at risk of problems with medication management. The PILL pharmacist called patients the week after hospital discharge to reconcile medications, assess adherence, and identify potentially inappropriate drugs. The pharmacist worked with each veteran's family and providers to resolve problems and increase support. To determine whether the intervention decreased acute care admissions, rehospitalizations, or deaths, we matched 1 unique control to each PILL patient by age, hospital location, length of stay, admitting service, and reason for admission. Logistic regression was performed to determine the OR and 95% CI of the outcomes. RESULTS: Patients were discharged on an average of 16 medications and with 4.4 medication changes. Overall, 61% of patients had clerical errors in the discharge summary, and potential clinical concerns were identified in over 75%. Veterans who received the intervention were 70% less likely than controls to have an acute care visit at 30 days postdischarge (7 vs 20 patients; OR = 0.30; 95% CI: 0.12-0.75). There was no difference in rates of hospital readmission or mortality. CONCLUSION: This pharmacist-led phone-based program was effective in decreasing acute care utilization within 30 days after hospital discharge.

Evaluation of oxygen prescription in relation to hospital admission rate in patients with chronic obstructive pulmonary disease.

BACKGROUND: Long term oxygen therapy (LTOT) has a strong evidence base in COPD patients with respiratory failure, but prescribing practices are recognized to need reform to ensure appropriate use and minimize costs. In the UK, since February 2006, all Home Oxygen prescription is issued by hospitals, making respiratory specialists totally in charge of home oxygen prescription. It has been widely noted that inappropriate home oxygen, often for intermittent use ("short burst"), is frequently prescribed in patients with COPD and related conditions with the intention to prevent hospital admissions outside of evidence based LTOT guidelines. We participated in a national Lung Improvement Project aimed at making LTOT use more evidence based. We utilised this unique opportunity of studying the effect of removal of oxygen from COPD patients (who did not meet LTOT criteria) on hospital admission rates. METHODS: Primary and secondary care data sources were used to identify patients with COPD in a single primary care trust who were admitted to hospital at least once due to COPD between April 2007 and November 2010. Admission rates were compared between LTOT users and non-users, adjusted for age and COPD severity. LTOT users were further studied for predictors of admission in those appropriately or inappropriately given oxygen according to NICE guidance, and for admissions before and after oxygen receipt, adjusting further for co-morbidity. Mortality and economic analyses were also conducted. RESULTS: Readmission was more likely in LTOT users (3.18 v 1.67 per patient, p<0.001) after adjustment for FEV1 and age by multiple regression. When stratifying by appropriateness of LTOT prescription, adjusting also for Charlson index and other covariates, FEV1 predicted admission in appropriate users but there were no predictors in inappropriate users. In longitudinal analyses admission rates did not differ either side of oxygen prescription in appropriate or inappropriate LTOT users. Specialist assessment resulted in cost savings due to reduced use of oxygen. CONCLUSIONS: Admission to hospital is more likely in LTOT users, independent of COPD severity. Oxygen use outside NICE guidance does not appear to prevent admissions.