Crystal structures of CDC21-1 inteins from hyperthermophilic archaea reveal the selection mechanism for the highly conserved homing endonuclease insertion site

Extremophiles - Self-splicing inteins are mobile genetic elements invading host genes via nested homing endonuclease (HEN) domains. All HEN domains residing within inteins are inserted at a highly...     

Epitope specificity of two anti‐morphine monoclonal antibodies: In vitro and in silico studies

Monoclonal antibodies (mAbs) against morphine are important in the development of immunotherapeutic and diagnostic methods for the treatment and prevention of drug addiction. By the surface plasmon resonance (SPR) and enzyme immunoassay techniques, we characterized two previously obtained mAbs 3K11 and 6G1 and showed their ability to recognize free morphine and morphine‐containing antigens in different ways because of the epitope specificity thereof. Using the defined amino acid sequences, we obtained three‐dimensional models of the variable regions of Fab fragments of these antibodies and compared them with the known sequence and spatial structure of the anti‐morphine antibody 9B1. Docking simulations are performed to obtain models of the antibodies complexes with morphine. Differences in the models of 3K11 and 6G1 complexes with morphine correlate with their experimentally detected epitope specificity. The results, in particular, can be used for the structure‐based design of the corresponding humanized antibodies. According to our modeling and docking results, the very different modes of morphine binding to mAbs 3K11 and 6G1 are qualitatively similar to those previously reported for cocaine and two anti‐cocaine antibodies. Thus, the obtained structural information brings more insight into the hapten recognition diversity.     

Human IL-36RA production in <Emphasis Type="Italic">Escherichia coli</Emphasis> with coexpression of <Emphasis Type="Italic">E. coli</Emphasis> methionine aminopeptidase. I. Comparison of IL-36RA production in different strains

Generalized pustular psoriasis (GPP) is a rare, sometimes lethal, form of psoriasis caused by series of mutations in the interleukin-36 receptor antagonist (IL-36RA) gene associated with its reduced expression or activity. Administration of exogenous IL-36RA can be a potent therapeutic approach to treating GPP and other forms of psoriasis. Since cleavage of the starting N-formylmethionine residue from the N-terminal end is needed for full biological activity of IL-36RA, we have developed a technique for producing IL-36RA lacking N-formylmethionine residue in E. coli. We have created a series of plasmids carrying the E. coli methionine aminopeptidase (MAP) gene under the control of different promoters for coexpression of IL-36RA and MAP and tested their effect on IL-36RA production. The highest production of IL-36RA with <3% of unprocessed molecules with uncleaved N-terminal formylmethionine residue has been shown for E. coli strain carrying the MAP gene under the control of arabinose-inducible promoter.