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  • Journal article

  • Three-dimensional envelope and subunit interactions of the plastid-encoded RNA polymerase from Sinapis alba

    • Authors : Ruedas, Rémi (2023)

  • RNA polymerases (RNAPs) involved in gene transcription are found in all living organisms with degrees of complexity ranging from single polypeptide chains to multimeric enzymes. In the chloroplasts, the nuclear-encoded RNA polymerase and the plastid-encoded RNA polymerase (PEP) are both involved in the selective transcription of the plastid genome. The PEP is a prokaryotic-type multimeric RNAP found in different states depending on light stimuli ...
  • Journal article

  • Filter paper disks as a matrix for manipulation of recombinant proteins

    • Authors : Ball, Eric H. (2023)

  • AbstractF ilter paper provides an excellent matrix for retention of proteins containing a cellulose binding domain. To use this capability for manipulating recombinant fusion proteins, binding and elution parameters were explored and procedures developed for small scale purification, modification and assay. Proteins were tagged with the cellulose binding domain from the C thermocellum CipB gene via a cleavable linker. Filter paper disks of 6mm&...
  • Journal article

  • Selenocyanate Derived Se-Incorporation into the Nitrogenase Fe Protein Cluster

    • Authors : Buscagan, Trixia M. (2023)

  • The nitrogenase Fe protein mediates ATP-dependent electron transfer to the nitrogenase MoFe protein during nitrogen fixation, in addition to catalyzing MoFe protein independent substrate (CO2) reduction and facilitating MoFe protein metallocluster biosynthesis. The precise role(s) of the Fe protein Fe4S4 cluster in some of these processes remains ill-defined. Herein, we report crystallographic data demonstrating ATP-dependent chalcogenide exchange at the Fe4S4 clus...
  • Journal article

  • The role of glutathione in periplasmic redox homeostasis and oxidative protein folding in Escherichia coli

    • Authors : Knoke, Lisa R. (2023)

  • The thiol redox balance in the periplasm of E. coli depends on the DsbA/B pair for oxidative 14 power and the DsbC/D system as its complement for isomerization of non-native disulfides. 15 While the standard redox potentials of those systems are known, the in vivo redox potential 16 imposed onto protein thiol disulfide pairs in the periplasm remains unknown. Here, we used 17 genetically encoded redox probes (roGFP2 ...
  • Journal article

  • Sulfotyrosine, an interaction specificity determinant for extracellular protein-protein interactions

    • Authors : Stewart, Valley (2023)

  • Tyrosine sulfation, a post-translational modification, can enhance and often determine protein-protein interaction specificity. Sulfotyrosyl residues (sTyr) are formed by tyrosylprotein sulfotransferase during maturation in the golgi apparatus, and most often occur singly or as a cluster of two or three sTyr within a six-residue span. With both negative charge and aromatic character, sTyr enables numerous atomic contacts as visualized in binding interfac...
  • Journal Article

  • Label-free Quantification of Host-Cell Protein Impurity in a Recombinant Hemoglobin Reference Material

    • Authors : Henrion, André (2023)

  • Quantitative analysis depends on pure-substance primary calibrators with known mass fractions of impurity. Here, label-free quantification (LFQ) is being evaluated as a readily available, reliable method for determining the mass fraction of host-cell proteins (HCPs) in bioengineered proteins. For example, hemoglobin-A2 (HbA2) is being used as obtained through overexpression in E.coli. Two different materials had been produced: natural, and U-15N-labeled HbA2. For quantification&...
  • Journal article

  • Ambient temperature crystal structure of Escherichia coli CyaY protein displays alternate conformation

    • Authors : Shafiei, Alaleh (2023)

  • Frataxin is a 23 KDa mitochondrial iron-binding protein that is involved in biogenesis of iron sulfur clusters. A deficiency in frataxin leads to Friedreich's ataxia, a progressive neurodegenerative disorder. The bacterial ortholog of eukaryotic mitochondrial frataxin, CyaY, is thought to play a role in iron sulfur cluster assembly as an iron supplier, making it an important target for study. Here, we present the first ambient tem...
  • Journal article

  • A database of accurate electrophoretic migration patterns for human proteins in cell lines

    • Authors : Mylonas, Roman (2023)

  • Native molecular weight (MW) is one of the defining features of proteins. Denaturing gel electrophoresis (SDS-PAGE) is a very popular technique for separating proteins and determining their MW. Coupled with antibody-based detection, SDS-PAGE is widely applied for protein identification and quantitation. Yet, electrophoresis is poorly reproducible and the MWs obtained are often inaccurate. This hampers antibody validation and negatively impacts the reliability of western...