Endonuclease I is a 149 amino acid protein of bacteriophage T7 that is a Holliday junction-resolving enzyme, i.e. a four-way junction-selective nuclease. We have performed a systematic mutagenesis study of this protein, whereby all acidic amino acids have been individually replaced by other residues, mainly alanine. Out of 21 acidic residues, five (Glu20, Glu35, Glu65, Asp55 and Asp74) are essential. Replacement of these residues by other amino acids leads to a protein that is inactive in the cleavage of DNA junctions, but which nevertheless binds selectively to DNA junctions. The remaining 16 acidic residues can be replaced without loss of activity. The five critical amino acids are located within one section of the primary sequence. It is rather likely that their function is to bind one or more metal ions that coordinate the water molecule that brings about hydrolysis of the phosphodiester bond. We have also constructed a mutant of endonuclease I that lacks nine amino acids (six of which are arginine or lysine) at the C-terminus. Unlike the acidic point mutants, the C-terminal truncation is unable to bind to DNA junctions. It is therefore likely that the basic C-terminus is an important element in binding to the DNA junction.
Two-component water assisted injection moulding (2K-WIT) is a new processing technology, fulfilling the high requirements of the automotive industry on media ducts on the one hand and offering a wide processing window and lower overall material costs on the other hand. The fundamental idea behind this technology is to split the requirements: mechanical strength, heat resistance, media resistance and channel surface quality into two groups. Each requirement group is then handled by a material which is tailored to fulfil these kinds of requirements. Schulamid 66 GF 30 2K WIT is used as the external skin material to achieve mechanical strength and temperature resistance. The inner wall surface is made out of polypropylene Polyfort FPP FX 2020 E, which offers perfect channel surface quality and ideal media resistance.
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