Effect of ampicillin-sulbactam on clinical capillary zone electrophoresis of serum proteins

Background: Capillary zone electrophoresis (CZE) is a well-accepted automated method used to separate serum proteins and detect monoclonal components. CZE uses ultraviolet detection at 214nm to directly quantify proteins via peptide bonds. Any substance that absorbs at 214nm and is present in serum can potentially interfere with CZE analysis. This has been reported for radio-contrast media and antibiotics. Methods: Here we describe a peak on the anode side of the α2-globulin fraction caused by the antibiotic ampicillin-sulbactam (Unacid®). Results and conclusions: Extra peaks that can be misinterpreted as monoclonal components can be present in almost all electrophoretic fractions of CZE. Immunosubtraction or immunofixation is always required to rule out these conditions. Clin Chem Lab Med 2008;46:1468-

Die quantitative Analyse von Markerproteinen im Urin Quantitative analysis of marker proteins in urine

Die Analyse von spezifischen Proteinen im zweiten Morgenurin, bezogen auf den Kreatiningehalt der Probe, erlaubt heute nicht nur den Nachweis oder den Ausschluss von Nierenerkrankungen, sondern darüber hinaus auch die Differenzierung und Verlaufskontrolle von Nephropathien. Störungen lassen sich aufgrund ihres Markerproteinprofils in solche mit hauptsächlich glomerulärem oder tubulärem Anteil und zusätzlich in weitere Untergruppen einteilen. Im Zusammenhang mit den Teststreifenresultaten kann die Quelle einer Blutung mit spezifischen Quotienten näher eingegrenzt und Kontaminationen können von tatsächlichen renalen Proteinurien unterschieden werden. Eine Plausibilitätsprüfung und Interpretation der erhaltenen Ergebnisse ist unbedingt erforderlich. Da eineVielzahl von Regeln überprüft werden muss, ist eine Berechnung und Darstellung der Ergebnisse nur mit Hilfe von wissensbasierten Systemen in Kombination mit einer grafischen Befunddarstellung sinnvol

Silenced yeast chromatin is maintained by Sir2 in preference to permitting histone acetylations for efficient NER

Very little is currently known about how nucleotide excision repair (NER) functions at the ends of chromosomes. To examine this, we introduced the URA3 gene into either transcriptionally active or repressed subtelomeric regions of the yeast genome. This enabled us to examine the repair of ultraviolet (UV)-induced cyclobutane pyrimidine dimers (CPDs) in identical sequences under both circumstances. We found that NER is significantly more efficient in the non-repressed subtelomere than the repressed one. At the non-repressed subtelomere, UV radiation stimulates both histones H3 and H4 acetylation in a similar fashion to that seen at other regions of the yeast genome. These modifications occur regardless of the presence of the Sir2 histone deacetylase. On the other hand, at the repressed subtelomere, where repair is much less efficient, UV radiation is unable to stimulate histone H4 or H3 acetylation in the presence of Sir2. In the absence of Sir2 both of these UV-induced modifications are detected, resulting in a significant increase in NER efficiency in the region. Our experiments reveal that there are instances in the yeast genome where the maintenance of the existing chromatin structures dominates over the action of chromatin modifications associated with efficient NER

Methylated H3K4, a Transcription-Associated Histone Modification, Is Involved in the DNA Damage Response Pathway

Eukaryotic genomes are associated with a number of proteins such as histones that constitute chromatin. Post-translational histone modifications are associated with regulatory aspects executed by chromatin and all transactions on genomic DNA are dependent on them. Thus, it will be relevant to understand how histone modifications affect genome functions. Here we show that the mono ubiquitylation of histone H2B and the tri-methylation of histone H3 on lysine 4 (H3K4me3), both known for their involvement in transcription, are also important for a proper response of budding yeast cells to DNA damaging agents and the passage through S-phase. Cells that cannot methylate H3K4 display a defect in double-strand break (DSB) repair by non-homologous end joining. Furthermore, if such cells incur DNA damage or encounter a stress during replication, they very rapidly lose viability, underscoring the functional importance of the modification. Remarkably, the Set1p methyltransferase as well as the H3K4me3 mark become detectable on a newly created DSB. This recruitment of Set1p to the DSB is dependent on the presence of the RSC complex, arguing for a contribution in the ensuing DNA damage repair process. Taken together, our results demonstrate that Set1p and its substrate H3K4me3, which has been reported to be important for the transcription of active genes, also plays an important role in genome stability of yeast cells. Given the high degree of conservation for the methyltransferase and the histone mark in a broad variety of organisms, these results could have similar implications for genome stability mechanisms in vertebrate and mammalian cells

Rif1 maintains telomeres and mediates DNA repair by encasing DNA ends

In yeast, Rif1 is part of the telosome, where it inhibits telomerase and checkpoint signaling at chromosome ends. In mammalian cells, Rif1 is not telomeric, but it suppresses DNA end resection at chromosomal breaks, promoting repair by nonhomologous end joining (NHEJ). Here, we describe crystal structures for the uncharacterized and conserved ∼125-kDa N-terminal domain of Rif1 from Saccharomyces cerevisiae (Rif1-NTD), revealing an α-helical fold shaped like a shepherd's crook. We identify a high-affinity DNA-binding site in the Rif1-NTD that fully encases DNA as a head-to-tail dimer. Engagement of the Rif1-NTD with telomeres proved essential for checkpoint control and telomere length regulation. Unexpectedly, Rif1-NTD also promoted NHEJ at DNA breaks in yeast, revealing a conserved role of Rif1 in DNA repair. We propose that tight associations between the Rif1-NTD and DNA gate access of processing factors to DNA ends, enabling Rif1 to mediate diverse telomere maintenance and DNA repair functions

DNA Damage during G2 Phase Does Not Affect Cell Cycle Progression of the Green Alga Scenedesmus quadricauda

DNA damage is a threat to genomic integrity in all living organisms. Plants and green algae are particularly susceptible to DNA damage especially that caused by UV light, due to their light dependency for photosynthesis. For survival of a plant, and other eukaryotic cells, it is essential for an organism to continuously check the integrity of its genetic material and, when damaged, to repair it immediately. Cells therefore utilize a DNA damage response pathway that is responsible for sensing, reacting to and repairing damaged DNA. We have studied the effect of 5-fluorodeoxyuridine, zeocin, caffeine and combinations of these on the cell cycle of the green alga Scenedesmus quadricauda. The cells delayed S phase and underwent a permanent G2 phase block if DNA metabolism was affected prior to S phase; the G2 phase block imposed by zeocin was partially abolished by caffeine. No cell cycle block was observed if the treatment with zeocin occurred in G2 phase and the cells divided normally. CDKA and CDKB kinases regulate mitosis in S. quadricauda; their kinase activities were inhibited by Wee1. CDKA, CDKB protein levels were stabilized in the presence of zeocin. In contrast, the protein level of Wee1 was unaffected by DNA perturbing treatments. Wee1 therefore does not appear to be involved in the DNA damage response in S. quadricauda. Our results imply a specific reaction to DNA damage in S. quadricauda, with no cell cycle arrest, after experiencing DNA damage during G2 phase

RNAi Screening Implicates a SKN-1-Dependent Transcriptional Response in Stress Resistance and Longevity Deriving from Translation Inhibition

Caenorhabditis elegans SKN-1 (ortholog of mammalian Nrf1/2/3) is critical for oxidative stress resistance and promotes longevity under reduced insulin/IGF-1-like signaling (IIS), dietary restriction (DR), and normal conditions. SKN-1 inducibly activates genes involved in detoxification, protein homeostasis, and other functions in response to stress. Here we used genome-scale RNA interference (RNAi) screening to identify mechanisms that prevent inappropriate SKN-1 target gene expression under non-stressed conditions. We identified 41 genes for which knockdown leads to activation of a SKN-1 target gene (gcs-1) through skn-1-dependent or other mechanisms. These genes correspond to multiple cellular processes, including mRNA translation. Inhibition of translation is known to increase longevity and stress resistance and may be important for DR-induced lifespan extension. One model postulates that these effects derive from reduced energy needs, but various observations suggest that specific longevity pathways are involved. Here we show that translation initiation factor RNAi robustly induces SKN-1 target gene transcription and confers skn-1-dependent oxidative stress resistance. The accompanying increases in longevity are mediated largely through the activities of SKN-1 and the transcription factor DAF-16 (FOXO), which is required for longevity that derives from reduced IIS. Our results indicate that the SKN-1 detoxification gene network monitors various metabolic and regulatory processes. Interference with one of these processes, translation initiation, leads to a transcriptional response whereby SKN-1 promotes stress resistance and functions together with DAF-16 to extend lifespan. This stress response may be beneficial for coping with situations that are associated with reduced protein synthesis

Effect of ampicillin-sulbactam on clinical capillary zone electrophoresis of serum proteins.

BACKGROUND: Capillary zone electrophoresis (CZE) is a well-accepted automated method used to separate serum proteins and detect monoclonal components. CZE uses ultraviolet detection at 214 nm to directly quantify proteins via peptide bonds. Any substance that absorbs at 214 nm and is present in serum can potentially interfere with CZE analysis. This has been reported for radio-contrast media and antibiotics. METHODS: Here we describe a peak on the anode side of the alpha(2)-globulin fraction caused by the antibiotic ampicillin-sulbactam (Unacid). RESULTS AND CONCLUSIONS: Extra peaks that can be misinterpreted as monoclonal components can be present in almost all electrophoretic fractions of CZE. Immunosubtraction or immunofixation is always required to rule out these conditions