N-acetyl glutamate kinase(NAGK)is a key enzyme in the synthesis of L-arginine,and L-arginine-sensitive NAGK typically has hexameric architecture.Defining the relationship between this architecture and L-arginine inhibition can provide a foundation to identify the key amino acids involved in the allosteric regulation network of L-arginine.In the present study,the key amino acids in the N-terminal helix(N-helix)of Corynebacterium glutamicum(Cg)NAGK required for hexamer formation were determined using structural homology modeling and site-directed mutagenesis.It was also verified that hexameric architecture is required for the positive cooperativity of inhibition by L-arginine and for efficient catalysis,but that it is not the determinant of inhibition by L-arginine.Monomeric mutants retained a similar sensitivity to L-arginine as the hexameric form,indicating that monomers contain an independent,sensitive signal transduction network of L-arginine to mediate allosteric regulation.Mutation studies of CgNAGKs also revealed that amino acid residues 18-23 of the N-helix are required for inhibition by Larginine,and that E19 may be an essential amino acid influencing the apparent affinity of L-arginine.Collectively,these studies may illuminate the basic mechanism of metabolic homeostasis of C.glutamicum.