PubMed | University Of Je Purkinje, CAS Institute of Microbiology, Institute of Biotechnology CAS, Czech Institute of Chemical Process Fundamentals and Leibniz Institute of Polymer Research
Type: Journal Article | Journal: Macromolecular bioscience | Year: 2016
For the design of a biohybrid structure as a ligand-tailored drug delivery system (DDS), it is highly sophisticated to fabricate a DDS based on smoothly controllable conjugation steps. This article reports on the synthesis and the characterization of biohybrid conjugates based on noncovalent conjugation between a multivalent biotinylated and PEGylated poly(amido amine) (PAMAM) dendrimer and a tetrameric streptavidin-small protein binding scaffold. This protein binding scaffold (SA-ABDwt) possesses nM affinity toward human serum albumin (HSA). Thus, well-defined biohybrid structures, finalized by binding of one or two HSA molecules, are available at each conjugation step in a controlled molar ratio. Overall, these biohybrid assemblies can be used for (i) a controlled modification of dendrimers with the HSA molecules to increase their blood-circulation half-life and passive accumulation in tumor; (ii) rendering dendrimers a specific affinity to various ligands based on mutated ABD domain, thus replacing tedious dendrimer-antibody covalent coupling and purification procedures.
PubMed | Czech Institute of Macromolecular Chemical, Institute of Biotechnology CAS, Charles University and CAS Institute of Microbiology
Type: Journal Article | Journal: Acta crystallographica. Section F, Structural biology communications | Year: 2016
Nepenthesins are aspartic proteases secreted by carnivorous pitcher plants of the genus Nepenthes. They significantly differ in sequence from other plant aspartic proteases. This difference, which provides more cysteine residues in the structure of nepenthesins, may contribute to their unique stability profile. Recombinantly produced nepenthesin 1 (rNep1) from N. gracilis in complex with pepstatin A was crystallized under two different crystallization conditions using a newly formulated low-pH crystallization screen. The diffraction data were processed to 2.9 and 2.8 resolution, respectively. The crystals belonged to space group P212121, with unit-cell parameters a = 86.63, b = 95.90, c = 105.40, = = = 90 and a = 86.28, b = 97.22, c = 103.78, = = = 90, respectively. Matthews coefficient and solvent-content calculations suggest the presence of two molecules of rNep1 in the asymmetric unit. Here, the details of the crystallization experiment and analysis of the X-ray data are reported.
PubMed | Novozymes AS, Institute of Biotechnology CAS and Institute of Chemical Technology Prague
Type: Journal Article | Journal: PloS one | Year: 2016
The single-strand-specific S1 nuclease from Aspergillus oryzae is an archetypal enzyme of the S1-P1 family of nucleases with a widespread use for biochemical analyses of nucleic acids. We present the first X-ray structure of this nuclease along with a thorough analysis of the reaction and inhibition mechanisms and of its properties responsible for identification and binding of ligands. Seven structures of S1 nuclease, six of which are complexes with products and inhibitors, and characterization of catalytic properties of a wild type and mutants reveal unknown attributes of the S1-P1 family. The active site can bind phosphate, nucleosides, and nucleotides in several distinguished ways. The nucleoside binding site accepts bases in two binding modes-shallow and deep. It can also undergo remodeling and so adapt to different ligands. The amino acid residue Asp65 is critical for activity while Asn154 secures interaction with the sugar moiety, and Lys68 is involved in interactions with the phosphate and sugar moieties of ligands. An additional nucleobase binding site was identified on the surface, which explains the absence of the Tyr site known from P1 nuclease. For the first time ternary complexes with ligands enable modeling of ssDNA binding in the active site cleft. Interpretation of the results in the context of the whole S1-P1 nuclease family significantly broadens our knowledge regarding ligand interaction modes and the strategies of adjustment of the enzyme surface and binding sites to achieve particular specificity.
PubMed | Institute of Physiology CAS, Institute of Experimental Medicine CAS, Institute of Biotechnology CAS and University of Iowa
Type: | Journal: Molecular neurobiology | Year: 2016
The programming of cell fate by transcription factors requires precise regulation of their time and level of expression. The LIM-homeodomain transcription factor Islet1 (Isl1) is involved in cell-fate specification of motor neurons, and it may play a similar role in the inner ear. In order to study its role in the regulation of vestibulo-motor development, we investigated a transgenic mouse expressing Isl1 under the Pax2 promoter control (Tg
PubMed | Max Planck Institute of Molecular Cell Biology and Genetics and Institute of Biotechnology CAS
Type: Journal Article | Journal: Current biology : CB | Year: 2016
Microtubules nucleated from an organizing center grow radially in all directions. A new study shows that, to organize those microtubules into arrays of parallel bundles, the kinesin-14 Cik1-Kar3 guides growing microtubule tips along pre-existing microtubules.
Biedermannova L.,Institute of Biotechnology CAS |
Schneider B.,Institute of Biotechnology CAS
Biochimica et Biophysica Acta - General Subjects | Year: 2016
Background Most biological processes involve water, and the interactions of biomolecules with water affect their structure, function and dynamics. Scope of review This review summarizes the current knowledge of protein and nucleic acid interactions with water, with a special focus on the biomolecular hydration layer. Recent developments in both experimental and computational methods that can be applied to the study of hydration structure and dynamics are reviewed, including software tools for the prediction and characterization of hydration layer properties. Major conclusions In the last decade, important advances have been made in our understanding of the factors that determine how biomolecules and their aqueous environment influence each other. Both experimental and computational methods contributed to the gradually emerging consensus picture of biomolecular hydration. General significance An improved knowledge of the structural and thermodynamic properties of the hydration layer will enable a detailed understanding of the various biological processes in which it is involved, with implications for a wide range of applications, including protein-structure prediction and structure-based drug design. © 2016 Elsevier B.V. All rights reserved.