Transglutaminases (EC 2.3.2.13) form an enzyme family that catalyzes the formation of isopeptide bonds between the γ-carboxamide group of glutamine and the ε-amine group of lysine residues of peptides and proteins. Other primary amines can be accepted in place of lysine. Because of their important physiological and pathophysiological functions, transglutaminases have been studied for 60 years. However, the substrate preferences of this enzyme class remain largely elusive. In this study, we used focused combinatorial libraries of 400 peptides to investigate the influence of the amino acids adjacent to the glutamine and lysine residues on the catalysis of isopeptide bond formation by microbial transglutaminase. Using the peptide microarray technology we found a strong positive influence of hydrophobic and basic amino acids, especially arginine, tyrosine, and leucine. Several tripeptide substrates were synthesized, and enzymatic kinetic parameters were determined both by microarray analysis and in solution.

L-Asparaginase is a protein produced by Escherichia coli and exists in two types, A and B. Only type B shows a specific antitumor activity to acute lymphatic leukemia. A new recombinant E. coli BL21Gold (DE3) pET11a-ansB was used to produce L-asparaginase B in higher quantities. To enhance the purity and preserve the activity of L-asparaginase B, crystallization is a promising process. Based on available patents, first experiments on L-asparaginase B crystallization were performed. Recombinant L-asparaginase B was successfully and reproducibly crystallized in two modifications. Up to now, it is not clear in which respect these two modifications of crystalline L-asparaginase B differ. It is part of the current study to investigate the nature of protein crystals in detail. The investigations on lysozyme crystals are well in agreement with the literature.

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