Oscillatory metabolism of Saccharomyces cerevisiae: an overview of mechanisms and models

The budding yeast Saccharomyces cerevisiae displays steady oscillations in continuous cultures under certain conditions. Oscillatory responses are important both metabolically and in process applications. Although much information has become available, a definitive theory to explain and model these oscillations is yet to be formulated. Models of oscillatory cultivation have focussed primarily either on intracellular reactions or on transport processes coupled to substantially lumped intracellular kinetics. This review discusses the development of the models and the directions they provide for a comprehensive model of oscillatory metabolism.     

The diagenetic fate of collagen as revealed by analytical pyrolysis of fossil fish scales from deep time

The mechanism of protein degradation has remained a topic of debate (specifically concerning their preservation in deep time), which has recently been invigorated due to multiple published reports of preservation ranging from Miocene to the Triassic that potentially challenge the convention that protein preservation beyond the Cenozoic is extremely uncommon or is expected to be absent altogether, and thus have attracted skepticism. In this paper, we analyze fossil fish scales from the Cretaceous, Jurassic, and Triassic using comprehensive pyrolysis gas chromatography coupled with time-of-flight mass spectrometry and compare the pyrolytic products so obtained with a well-preserved fish scale from Late Pliocene, in an attempt to better understand the effects of diagenesis on protein degradation at the molecular level through deep time. We find that the Pliocene fish scale displays a large number of N-bearing pyrolytic products, including abundant substituted cyclic 2,5-diketopiperazines (2,5-DKPs) which are diagnostic products of peptide and amino acid pyrolysis. We identify N-bearing compounds in the Mesozoic fish scales—however, among the 2,5-DKPs that were identified in the Pliocene scale, only diketodipyrrole (or cyclo (Pyr-Pyr)) is present in the Mesozoic scales. We discuss the implications of N-bearing pyrolytic products with emphasis on 2,5-DKPs in geological samples and conclude that the discrepancy in abundance and variety of N-bearing products between Pliocene and Mesozoic scales indicates that the protein component in the latter has been extensively diagenetically altered, while a suite of DKPs such as in the former would imply stronger evidence to indicate preservation of protein. We conclude that analytical pyrolysis is an effective tool for detecting preservation of intact proteins, as well as for providing insights into their degradation mechanisms, and can potentially be utilized to assign proteinaceous origin to a fossil sample of unknown affinity.     

Studies on the binding of d-α-tocopherol to rat liver nuclei

The present study describes the nature and characteristics of the intranuclear binding sites of [³H]d-α-tocopherol in rat liver. When radioactively labeled d-α-tocopherol was intravenously administered to rats, approximately 55% of the nuclear radioactivity was associated with an intranuclear nucleoprotein complex. This complex, which was extractable by high concentrations of NaCl, was characterized by equilibrium density ultracentrifugation on a 30 to 60% linear sucrose gradient. About 50% of the high-salt-extracted radioactivity was coprecipitable with macromolecules by 10% ice-cold trichloroacetic acid (TCA). This TCA-precipitable radioactivity was completely ethanol soluble. Alkaline conditions favored the solubilization of the vitamin-receptor complex. Among various enzymes tested, only Pronase and trypsin were capable of dissociating the vitamin-receptor complex. Both ionic (sodium dodecyl sulfate) and nonionic (Triton X-100) detergents solubilized α-tocopherol from the nuclei and concomitantly released some of the associated macromolecules. In addition, treatment of nuclei with low concentrations of Triton X-100 showed that about 30% of the nuclear bound α-tocopherol is associated with inner core sites in the nucleoprotein complex with very high affinity for the vitamin. Dissociation of the nucleoprotein complex (chromatin) by high-salt solubilization and subsequent partial reassociation of the components by salting out procedures revealed the high affinity association of α-tocopherol with the reconstituted DNA-protein complex. Subfractionation of this complex further revealed that α-tocopherol is predominantly associated with the fraction containing phenol-soluble nonhistone proteins having a high affinity for DNA. In vitro binding studies also showed that there are specific saturable binding sites for d-α-tocopherol in rat liver nuclei.     

Identification of highly potent competence stimulating peptide-based quorum sensing activators in Streptococcus mutans through the utilization of N-methyl and reverse alanine scanning

Quorum sensing (QS) controls the pathogenic behavior of Streptococcus mutans, a primary cause of dental caries. S. mutans uses the competence stimulating peptide (CSP) to control mutacin production, a bacteriocin utilized by S. mutans to outcompete different commensal bacteria in mixed biofilm environments. In this study, we performed an N-methyl scan of an 18-CSP-based scaffold lacking the first two amino acid residues that were shown to be dispensable, to gain important mechanistic insight as to the role of backbone amide protons in the interaction between CSP and the ComD receptor. We then utilized the reverse alanine approach to develop CSP-based analogs with enhanced activities. The two most potent analogs were found to induce bacteriocin production at sub-nanomolar concentration using an interspecies inhibition assay. Overall, our analysis revealed that the 18-CSP sequence is not optimized and can be improved by replacement of multiple positions with alanine. Our results further suggest that the hydrophobic residues in S. mutans 18-CSP are involved in both receptor binding and activation.     

Phase 1 trial of the oral AKT inhibitor MK-2206 plus carboplatin/paclitaxel, docetaxel, or erlotinib in patients with advanced solid tumors

Background

Inhibition of AKT with MK-2206 has demonstrated synergism with anticancer agents. This phase 1 study assessed the MTD, DLTs, PK, and efficacy of MK-2206 in combination with cytotoxic and targeted therapies.

Methods

Advanced solid tumor patients received oral MK-2206 45 or 60 mg (QOD) with either carboplatin (AUC 6.0) and paclitaxel 200 mg/m² (arm 1), docetaxel 75 mg/m² (arm 2), or erlotinib 100 or 150 mg daily (arm 3); alternative schedules of MK-2206 135-200 mg QW or 90-250 mg Q3W were also tested.

Results

MTD of MK-2206 (N?=?72) was 45 mg QOD or 200 mg Q3W (arm 1); MAD was 200 mg Q3W (arm 2) and 135 mg QW (arm 3). DLTs included skin rash (arms 1, 3), febrile neutropenia (QOD, arms 1, 2), tinnitus (Q3W, arm 2), and stomatitis (QOD, arm 3). Common drug-related toxicities included fatigue (68%), nausea (49%), and rash (47%). Two patients with squamous cell carcinoma of the head and neck (arm 1; Q3W) demonstrated a complete and partial response (PR); additional PRs were observed in patients (1 each) with melanoma, endometrial, neuroendocrine prostate, NSCLC, and cervical cancers. Six patients had stable disease ≥6 months.

Conclusion

MK-2206 plus carboplatin and paclitaxel, docetaxel, or erlotinib was well-tolerated, with early evidence of antitumor activity.

Trial registration

ClinicalTrials.gov: NCT00848718.     

Current trends in large‐scale viral surveillance methods in mosquitoes

Vector‐borne and zoonotic infectious diseases are serious public health concerns that affect approximately half of the world's population. In particular, arthropod‐borne viruses (arboviruses) have contributed to more mortality and morbidity worldwide with the emergence of dengue, chikungunya, yellow fever, and Zika virus diseases. The infections have scaled up due to urbanization, globalization, and international mobility. Traditionally, the spread of mosquito‐borne viral diseases to humans was considered a low health priority concern. However, their categorization as emerging infectious diseases and public health emergencies of international concern has heightened the attention given by the government, academia, research, and industry for the development of timely, cost‐efficient, and sustainable solutions. The urgency has increased in the wake of global climate change. The focus on effective interventions includes epidemiological monitoring, vector control measures, molecular diagnostics, vaccines, and environmental determinants. In this review, we discuss the etiology and predisposition of mosquito‐borne viruses that are detrimental to public health and economically damaging when disseminated as epidemics. We focus on the large‐scale virus surveillance methods with special reference to innovations and interventions in molecular detection science and technologies that include viral nucleic acid isolation, polymerase chain reaction (PCR)‐based diagnostics, and high‐throughput sequencing technologies. In addition, we discuss the development of a viral RNA extraction and PCR‐based diagnostic kit (Invirustech) that can extract viral RNA from mosquitoes with verified applications in PCR‐based molecular diagnostics of Pan‐flavivirus.     

Simultaneous detection of potato viruses Y and X by DAC-ELISA using polyclonal antibodies raised against fused coat proteins expressed in Escherichia coli

Polyclonal antibodies were raised against the bacterial expressed fused coat proteins (CPs) of Potato virus Y (PVY) and Potato virus X (PVX). Truncated CP sequences of PVY (~246 bp) and PVX (~243 bp) were amplified by PCR, cloned into T&A cloning vector and subsequently mobilized in a protein expression vector pET-28b (+). The recombinant CP was expressed as a fusion protein (~20 kDa) with His-tag and purified from E. coli BL21 (DE3) using His-Bind resin. The specificity of the recombinant protein was confirmed by Western blot using previously made polyclonal antibodies against each virus. Polyclonal antibodies developed against the fused CPs in rabbit detected natural infection of PVY and PVX in potato leaf samples collected from IARI experimental farm, by direct antigen-coated enzyme-linked immunosorbent assay (DAC-ELISA).     

Age-related responses of hepatic ATPases to starvation and cold stress in male garden lizards

Hepatic Na⁺-K⁺-ATPase and Mg²⁺-ATPase activities of male green lizards declined during the maturation phase (juvenile to 1-year-old) and stabilized thereafter. On the other hand, the Ca²⁺-ATPase activity of the liver declined during the later half of the life span (1-year-old to 2–4-year-old). Starvation stress induced a decline in hepatic Na⁺-K⁺-ATPase and Mg²⁺-ATPase activities of juvenile lizards and caused an increase in 1-year-old and 2–4-year-old counterparts. The Ca²⁺-ATPase activity declined only in starved 1-year-old lizards. Following cold stress, the hepatic Na⁺-K⁺-ATPase activity of juvenile lizards showed a higher degree of decline than 2–4-year-old counterparts. The Mg²⁺-ATPase activity declined in cold-stressed juvenile lizards, but the parameter was elevated in similarly treated 1-year-old lizards. On the other hand, the increase in Ca²⁺-ATPase activity in response to cold stress was confined only to 2–4-year-old lizards.