Chitinase A (ChiA) from the bacterium Serratia marcescens is a hydrolytic enzyme, which cleaves beta-1,4-glycosidic bonds of the natural biopolymer chitin to generate di-N-acetyl-chitobiose. The refined structure of ChiA at 1.55 A shows that residue Asp313, which is located near the catalytic proton donor residue Glu315, is found in two alternative conformations of equal occupancy. In addition, the structures of the cocrystallized mutant proteins D313A, E315Q, Y390F, and D391A with octa- or hexa-N-acetyl-glucosamine have been refined at high resolution and the interactions with the substrate have been characterized. The obtained results clearly show that the active site is a semiclosed tunnel. Upon binding, the enzyme bends and rotates the substrate in the vicinity of the scissile bond. Furthermore, the enzyme imposes a critical "chair" to "boat" conformational change on the sugar residue bound to the -1 subsite. According to our results, we suggest that residues Asp313 and Tyr390 along with Glu315 play a central role in the catalysis. We propose that after the protonation of the substrate glycosidic bond, Asp313 that interacts with Asp311 flips to its alternative position where it interacts with Glu315 thus forcing the substrate acetamido group of -1 sugar to rotate around the C2-N2 bond. As a result of these structural changes, the water molecule that is hydrogen-bonded to Tyr390 and the NH of the acetamido group is displaced to a position that allows the completion of hydrolysis. The presented results suggest a mechanism for ChiA that modifies the earlier proposed "substrate assisted" catalysis.

γH2AX has emerged in the last 20 years as a central player in the DDR (DNA damage response), with specificity for DSBs (double-strand breaks). Upon the generation of DSBs, γ-phosphorylation extends along megabase-long domains in chromatin, both sides of the damage. The significance of this mechanism is of great importance; it depicts a biological amplification mechanism where one DSB induces the γ-phosphorylation of thousands of H2AX molecules along megabaselong domains of chromatin, that are adjusted to the sites of DSBs. A sequential recruitment of signal transduction factors that interact to each other and become activated to further amplify the signal that will travel to the cytoplasm take place on the γ-phosphorylated chromatin. γ-phosphorylation is an early event in the DSB damage response, induced in all phases of the cell cycle, and participates in both DSB repair pathways, the HR (homologous recombination) and NHEJ (non-homologous end joining). Today, numerous studies support the notion that γH2AX functions as a guardian of the genome by preventing misrepaired DSB that increase the mutation load of the cells and may further lead to genome instability and carcinogenesis.

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