The Cryo-EM Structure of a Complete 30S Translation Initiation Complex from Escherichia coli

Formation of the 30S initiation complex (30S IC) is an important checkpoint in regulation of gene expression. The selection of mRNA, correct start codon, and the initiator fMet-tRNAfMet requires the presence of three initiation factors (IF1, IF2, IF3) of which IF3 and IF1 control the fidelity of the process, while IF2 recruits fMet-tRNAfMet. Here we present a cryo-EM reconstruction of the complete 30S IC, containing mRNA, fMet-tRNAfMet, IF1, IF2, and IF3. In the 30S IC, IF2 contacts IF1, the 30S subunit shoulder, and the CCA end of fMet-tRNAfMet, which occupies a novel P/I position (P/I1). The N-terminal domain of IF3 contacts the tRNA, whereas the C-terminal domain is bound to the platform of the 30S subunit. Binding of initiation factors and fMet-tRNAfMet induces a rotation of the head relative to the body of the 30S subunit, which is likely to prevail through 50S subunit joining until GTP hydrolysis and dissociation of IF2 take place. The structure provides insights into the mechanism of mRNA selection during translation initiation

Translation initiation factor IF1 of Bacillus stearothermophilus and Thermus thermophilus substitute for Escherichia coli IF1 in vivo and in vitro without a direct IF1-IF2 interaction.

Bacterial translation initiation factor IF1 is homologous to archaeal aIF1A and eukaryal eIF1A, which form a complex with their homologous IF2-like factors (aIF5B and eIF5B respectively) during initiation of protein synthesis. A similar IF1-IF2 interaction is assumed to occur in all bacteria and supported by cross-linking data and stabilization of the 30S-IF2 interaction by IF1. Here we compare Escherichia coli IF1 with thermophilic factors from Bacillus stearothermophilus and Thermus thermophilus. All three IF1s are structurally similar and functionally interchangeable in vivo and in vitro. However, the thermophilic factors do not stimulate ribosomal binding of IF2DeltaN, regardless of 30S subunits and IF2 origin. We conclude that an IF1-IF2 interaction is not universally conserved and is not essential for cell survival

Refractory septic shock: Efficacy and safety of very high doses of norepinephrine

The aim of Ms study was to evaluate the safety, efficacy, and effects of administration of very high doses of norepinephrine (>4 μg kg-1 min-1) in catecholamine-resistant septic shock. We reviewed the charts of all patients with nonresponding to commonly used norepinephrine doses (≤4 μg kg-1 min-1) septic shock from January 1999 to December 2002 in our Surgical Intensive Care Unit. All patients were treated with high norepinephrine doses (>4 μg kg-1 min-1), after initial resuscitation, so as to achieve a mean arterial pressure higher than or equal to 65 mmHg. During this 4-year period, 12 consecutive patients with catecholamine-resistant septic shock were included in our study. When compared with the values obtained prior to the administration of very high norepinephrine doses, the values of mean arterial pressure (p = 0.003) and systemic vascular resistance (p = 0.002) significantly increased after the administration of such doses, and additionally, lactate concentrations (p = 0.003) decreased. In contrast, no significant changes were observed regarding mean central venous pressure, pulmonary capillary wedge pressure, and pulmonary arterial pressure. Administration of high norepinephrine doses in our patients resulted in a survival rate of 33.4%. Management of catecholamine-resistant septic shock patients poses a challenging problem. Administration of very high norepinephrine doses is safe and effective and may improve survival of these patients with otherwise extremely high mortality rates. © 2006 Prous Science. All rights reserved

A simplified technique for translaryngeal tracheostomy (TLT). A preliminary report

Objective In this prospective observational study we present preliminary results of a modification of the translaryngeal tracheostomy technique that was introduced by Fantoni in 1997. The study was conducted in a five-bed surgical intensive care unit of a university teaching hospital. Patients and Methods The study included 14 consecutive surgical patients (8 men, 6 women) who underwent a modified translaryngeal tracheostomy in a 6-month period. In our modification of the technique, we keep the basic principle of the inside-to-outside approach of the Fantoni technique, and combine it with a blind needle insertion, as reported in the classic subcricoid retrograde intubation technique. The technique that we use involves two medical doctors and a nurse. Results Mean patient age was 68.9 years (range: 31-85 years) and mean APACHE II score was 15.8 (range: 6-31). Mean operative time for the procedure was 15.2 min (range: 11.5-22 min). Eight of the patients died during the postoperative course in the ICU from causes relevant to their surgical pathology. One patient survived to be discharged from the ICU but died of an acute myocardial infraction later in the same hospital stay. Five patients survived to be discharged from the hospital. Conclusions The modified translaryngeal tracheostomy seems to be as reliable and safe as the original technique. In addition, the modified technique is faster and can be performed without the use of an endoscope