title: Catalytic activity of Peptidase N is required for adaptation of Escherichia coli to nutritional downshift and high temperature stress.
abstract: Peptidase N (PepN), the sole M1 family member in Escherichia coli, displays broad substrate specificity and modulates stress responses: it lowers resistance to sodium salicylate (NaSal)-induced stress but is required during nutritional downshift and high temperature (NDHT) stress. The expression of PepN does not significantly change during different growth phases in LB or NaSal-induced stress; however, PepN amounts are lower during NDHT stress. To gain mechanistic insights on the roles of catalytic activity of PepN in modulating these two stress responses, alanine mutants of PepN replacing E264 (GAMEN motif) and E298 (HEXXH motif) were generated. There are no major structural changes between purified wild type (WT) and mutant proteins, which are catalytically inactive. Importantly, growth profiles of ΔpepN upon expression of WT or mutant proteins demonstrated the importance of catalytic activity during NDHT but not NaSal-induced stress. Further fluorescamine reactivity studies demonstrated that the catalytic activity of PepN is required to generate higher intracellular amounts of free N-terminal amino acids; consequently, the lower growth of ΔpepN during NDHT stress increases with high amounts of casamino acids. Together, this study sheds insights on the expression and functional roles of the catalytic activity of PepN during adaptation to NDHT stress.
authors: Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors: Manoj
authors: M
authors: Bhosale
authors: Anujith
authors: A
authors: Kumar
authors: Mrinmoy
authors: M
authors: Das
authors: Chetana
authors: C
authors: Bhaskarla
authors: Vikas
authors: V
authors: Agarwal
authors: Dipankar
authors: D
authors: Nandi
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000626:Aminopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology* / Q000528:radiation effects; D020869:Gene Expression Profiling; D006358:Hot Temperature; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D016415:Sequence Alignment; D013312:Stress, Physiological
chemical_list: D050505:Mutant Proteins; D000626:Aminopeptidases; C034113:peptidase N