THE ACTIVATION OF ESCHERICHIA-COLI ASPARTATE-TRANSCARBAMYLASE BY ATP - SPECIFIC INVOLVEMENT OF HELIX H2' AT THE HYDROPHOBIC INTERFACE BETWEEN THE 2 DOMAINS OF THE REGULATORY CHAINS

The regulatory chain of E. coli aspartate transcarbamylase (E.C. 2.1.3 .2) is folded into two domains. The allosteric domain harbours the reg ulatory site where the activator ATP and the inhibitors CTP and UTP bi nd competitively. The zinc domain ensures the contact with the catalyt ic chains. The interface between these two domains is hydrophobic, and involves the carboxy-terminal part of the helix H2' of the allosteric domain and several residues of the zinc domain. This structural featu re mediates the transmission of the ATP regulatory signal. In the pres ent work, site-directed mutagenesis and molecular modelling were used to investigate the role of specific amino acid residues in this proces s. The modifications of the hydrophobic core which are expected to alt er the position of helix H2' reduce or abolish the sensitivity of the enzyme to ATE The properties of the mutants and the results of modelli ng are fully consistent and suggest that a movement of helix H2' is pa rt of the mechanism of activation by ATP A model is proposed to accoun t for the transmission of the ATP signal from the regulatory site to t he interface between the regulatory and catalytic chains.