QUASI-ELASTIC LIGHT SCATTERING STUDIES OF THE KINETICS OF LYSOZYME DIMERIZATION

On a utilisé la technique de la diffusion de la lumière laser pour étudier les vitesses et le mécanisme de dimérisation du lysozyme en l'absence de substrat. Ces expériences ont été réalisées sur du lysozyme dans du KCl 0,15 molaire, a 20 °C , dans un domaine de pH de 3,0 à 7,0. Les concentrations en protéine ont varié entre 3,6 x 10-4 et 14,6 x 10-4 molaire. L'échantillon de pH = 3,0 (7,3 x 10-4 M) a présenté un Dw20 = (1,04 ± 0,02) x 10-6 cm2/s, correspondant bien à la valeur acceptée pour le coefficient de diffusion translationnel de la forme monomère du lysozyme. Les données de la diffusion ont indiqué que pour un pH 5,0-7,0, le lysozyme subit une transformation de configuration avant de prendre la forme dimère stable. Cela est mis en évidence par un minimum dans la valeur du coefficient de diffusion apparent au-dessous de ce qu'on attend pour le dimère. Une analyse détaillée des spectres observés indique qu'on peut discerner deux étapes dans la réaction. La plus lente de ces étapes correspond à la valeur de saut-T de Owen, Eyring et Cole (1969), qui pour un pH de 7,0 est de (2,0 ± 0,5) x 10-3 s. Les données présentes pour la réaction la plus lente montrent une absence semblable de dépendance avec la température pour pH = 7,0 comme l'ont trouvé Owen et Coll. Cependant, pour des pH de 5,0 et 6,0, il y a une augmentation du temps de relaxation avec une diminution de la concentration (au-dessous de 7 x 10-4 M de concentration de protéine) pour cette étape. D'après les données de la diffusion, il apparaît aussi une composante spectrale correspondant à une étape de réaction avec un temps de relaxation de (3,5 ± 1,0) x 10-5 s. La vitesse de ce processus plus rapide ne varie pas avec le pH ou la concentration en protéine dans les limites des incertitudes expérimentales. On a postulé un modèle pour le mécanisme de dimérisation en deux étapes faisant intervenir un changement de configuration et un processus bimoléculaire. Ce modèle est capable de donner un accord qualitatif avec les données observées bien qu'une nouvelle étude de l'auto-dissociation du lysozyme par ultracentrifugation suggère que pour le pH de 7,0, l'association soit plus élevée que dimère.The laser light scattering technique has been used to probe the rates and mechanism of the dimerization of lysozyme in the absence of substrates. These experiments were performed on lysozyme in 0.15 M KCl and 20 °C over a range of pH from 3.0 to 7.0. Protein concentration varied from 3.6 x 10-4 M to 14.6 x 10-4 M. The pH = 3.0 sample (7.3 x 10-4 M) showed that Dw20 = (1.04 ± .02) x 10-6 cm2/s, corresponding well to the accepted value of the translational diffusion coefficient for the monomeric form of lysozyme. The scattering data indicated that at pH 5.0-7.0, lysozyme undergoes a conformation change before assuming the stable dimer form. This is evidenced by a minimum in the value of the apparent diffusion coefficient below that expected for the dimer. Detailed analysis of the observed spectra indicates that two rate steps can be discerned. The slower of the two rates corresponds to the T-jump value of Owen, Eyring, and Cole, (1969), which at pH = 7.0 is (2.0 ± .5) x 10-3 s. The present data for the slower reaction show a similar lack of concentration dependence at pH = 7.0 as was found by Owen, et al. However, at pH = 5.0 and 6.0, there is an increase of the relaxation time with decreasing concentration (below 7 x 10-4 M protein concentration) for this step. Also evident from the scattering data is a spectral component corresponding to a rate process with a relaxation time of (3.5 ± 1.0) x 10-5 s. The rate of this faster process does not Vary with pH or protein concentration within the bounds of experimental uncertainty. A model for a two-step dimerization mechanism involving a conformation change and a bimolecular process has been postulated. This model is able to give qualitative agreement with the observed data although a reinvestigation of the lysozyme self-dissociation by equilibrium ultracentrifugation suggest that at pH = 7.0, the association may be higher than dimer

Antibiotic combination therapy against resistant bacterial infections: synergy, rejuvenation and resistance reduction.

Introduction: Anti-Microbial Resistance (AMR) is a pandemic which threatens modern medicine. There is a lack of effective drug treatment due to the slow pace, high cost and low achievable sales prices of new antibiotic monotherapies. A new hope comes in the shape of antibiotic combination therapy, which although used by mother nature, is under-explored and could provide the solution to AMR.Areas covered: We performed a search of Pubmed and Medline using the keywords "combination therapy", "antimicrobial resistance" for articles between 1930 and 2019, as supplemented with other relevant references to our knowledge. We will review the theoretical considerations for combination development and examine the existing and future clinical indications of combination therapies. We will discuss the potential of antibiotic combinations to provide therapeutic synergy, rejuvenating the effectiveness of old antibiotics to which the bacteria had developed resistance previously. We will examine the current thinking and evidence on resistance reduction using combination therapies, with a review on toxicity and drug-drug antagonism.Expert opinion: Antibiotic combination therapy, exploiting synergies, old-drug rejuvenation and resistance reduction could provide the solution to AMR. The number of pharmaceutical companies in this area is likely to expand, bringing promising combinations to the bedside, to save millions of lives worldwide

The haemodynamic effect of verapamil on portal hypertension in patients with postnecrotic cirrhosis

Phenolic Impregnated Carbon Ablator (PICA), invented in the mid 1990's, is a low-density ablative thermal protection material proven capable of meeting sample return mission needs from the moon, asteroids, comets and other unrestricted class V destinations as well as for Mars. Its low density and efficient performance characteristics have proven effective for use from Discovery to Flag-ship class missions. It is important that NASA maintain this thermal protection material capability and ensure its availability for future NASA use. The rayon based carbon precursor raw material used in PICA preform manufacturing has experienced multiple supply chain issues and required replacement and requalification at least twice in the past 25 years and a third substitution is now needed. The carbon precursor replacement challenge is twofold - the first involves finding a long-term replacement for the current rayon and the second is to assess its future availability periodically to ensure it is sustainable and be alerted if additional replacement efforts need to be initiated. This paper reviews current PICA sustainability activities to identify a rayon replacement and to establish that the capability of the new PICA derived from an alternative precursor is in family with previous versions