Ku is a heterodimer composed of p70 and p80, and is the regulatory subunit of DNA-dependent protein kinase. As a multifunctional DNA-binding protein complex, Ku plays important roles in DNA damage repair through non-homologous end joining and in V(D)J recombination. In addition, Ku has also been implicated in various biological functions including growth control, cell proliferation, cell cycle, chromosome maintenance, transcriptional regulation, apoptosis, and viral infection. In particular, using our Inverse Genomics (Immusol, Inc., San Diego, CA) platform technology, we recently identified Ku80 as an essential co-factor for human immunodeficiency virus replication. Although Ku has been studied extensively in the past years, its in-depth study as well as development as a drug target has been limited by conventional DNA-binding activity assay. Here we describe the development and applications of a nonradioactive DNA binding assay in the 96-well format. We show that this plate-formatted assay is more sensitive and allows for direct quantification when compared with an electrophoretic mobility shift assay. The establishment of this assay will not only facilitate structure and function studies on Ku, but also help the development of Ku protein or its DNA repair enzyme complex as a drug target.
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http://dx.doi.org/10.1089/adt.2004.2.483 | DOI Listing |
Biomol NMR Assign
January 2025
High Magnetic Field Laboratory, Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei, Anhui, China.
PhoCl is a photocleavable protein engineered from a green-to-red photoconvertible fluorescent protein by circular permutation, and has been used in various optogenetic applications including precise control of protein localization and activity in cells. Upon violet light illumination, PhoCl undergoes a β-elimination reaction to be cleaved at the chromophore, resulting in spontaneous dissociation into a large empty barrel and a small C-terminal peptide. However, the structural determinants and the mechanism of the PhoCl photocleavage remain elusive, hindering the further development of more robust photocleavable optogenetic tools.
View Article and Find Full Text PDFJ Clin Monit Comput
January 2025
Department of Anaesthesiology and Intensive Care, Bicetre hospital, Assistance Publique Hôpitaux de Paris (AP-HP), Le Kremlin Bicetre, France.
Intravenous fluid is administered during high-risk surgery to optimize stroke volume (SV). To assess ongoing need for fluids, the hemodynamic response to a fluid bolus is evaluated using a fluid challenge technique. The Acumen Assisted Fluid Management (AFM) system is a decision support tool designed to ease the application of fluid challenges and thus improve fluid administration during high-risk surgery.
View Article and Find Full Text PDFIntensive Care Med Exp
January 2025
Freie Universität Berlin and Humboldt-Universität Zu Berlin, Department of Anesthesiology and Intensive Care Medicine, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany.
Purpose: CytoSorb® (CS) adsorbent is a hemoadsorption filter for extracorporeal blood purification often integrated into continuous kidney replacement therapy (CKRT). It is primarily used in critically ill patients with sepsis and related conditions, including cytokine storms and systemic inflammatory responses. Up to now, there is no evidence nor recommendation for the use of CS filters in sepsis (22).
View Article and Find Full Text PDFTop Curr Chem (Cham)
January 2025
School of Pharmacy, Hangzhou Normal University, Hangzhou, 311121, Zhejiang, People's Republic of China.
BCP (bicyclo[1.1.1]pentane) is an ideal saturated carbon bioisostere, instead of the traditional benzene group, which has been extensively developed.
View Article and Find Full Text PDFJ Mammary Gland Biol Neoplasia
January 2025
Department of Histology and Embryology, Faculty of Medicine, Masaryk University, Brno, Czech Republic.
Fluorescent biosensors offer a powerful tool for tracking and quantifying protein activity in living systems with high temporospatial resolution. However, the expression of genetically encoded fluorescent proteins can interfere with endogenous signaling pathways, potentially leading to developmental and physiological abnormalities. The EKAREV-NLS mouse model, which carries a FRET-based biosensor for monitoring extracellular signal-regulated kinase (ERK) activity, has been widely utilized both in vivo and in vitro across various cell types and organs.
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