Activation by Allostery in Cell-Wall Remodeling by a Modular Membrane-Bound Lytic Transglycosylase from Pseudomonas aeruginosa

Abstract

SUMMARY Bacteria grow and divide without loss of cellular integrity. This accomplishment is notable, as a key component of their cell envelope is a surrounding glycopeptide polymer. In Gram-negative bacteria this polymer—the peptidoglycan—grows by the difference between concurrent synthesis and degradation. The regulation of the enzymatic ensemble for these activities is poorly understood. We report herein the structural basis for the control of one such enzyme, the lytic transglycosylase MltF of Pseudomonas aeruginosa. Its structure comprises two modules: an ABC-transporter-like regulatory module and a catalytic module. Occupancy of the regulatory module by peptidoglycan-derived muropeptides effects a dramatic and long distance (40 Å) conformational change, occurring over the entire protein structure, to open its active site for catalysis. This discovery of the molecular basis for the allosteric control of MltF catalysis is foundational to further study of MltF within the complex enzymatic orchestration of the dynamic peptidoglycan.

Lytic transglycosylases: concinnity in concision of the bacterial cell wall

“…MltF lacks function either in side-wall growth or in septum formation in both E. coli and P. aeruginosa (Scheurwater & Clarke, 2008; Lamers et al , 2015). Accordingly, its assignment may be for cell-wall excavation so as to enable the passage of the pilli, fimbriae, and other macromolecular complexes through the peptidoglycan (Dominguez-Gil et al , 2016). Structures of the apo form of P. aeruginosa MltF (PA3764), and its complexes with bulgecin, muropeptide, and various amino acids (cysteine, valine, leucine, and isoleucine) in its ABC-transporter domain were deposited by Thunnissen in 2015, but without accompanying publication (Madoori & Thunnissen, 2010).…”

“…Structures of the apo form of P. aeruginosa MltF (PA3764), and its complexes with bulgecin, muropeptide, and various amino acids (cysteine, valine, leucine, and isoleucine) in its ABC-transporter domain were deposited by Thunnissen in 2015, but without accompanying publication (Madoori & Thunnissen, 2010). Complementary structures of active and inactive P. aeruginosa MltF (Figure 5) in complex with NAG-anhNAM-pentapeptide, NAM-pentapeptide, tetrasaccharide and tetrapeptide, and tetrapeptide gave insight into its allosteric regulation (Dominguez-Gil et al , 2016). The structure of the N-terminal SBP_bac_3 regulatory module of MltF, comprising of two subdomains connected by a flexible linker, resembles that of an ABC-transporter domain.…”

“…Previously, GP144 was assigned as a Family 4 LT only on the basis of its bacteriophage origin (Domínguez-Gil et al , 2016). The distinction between Family 1 LTs and Family 4 LTs (such as E. coli bacteriophage lambda lysozyme) is evident from structure analysis of the catalytic domains.…”

“…Family 2 belongs to the non-G-type GH102 sub-family. The well-studied chicken-type (C-type) lysozyme belongs to the GH22 sub-family (Gloster & Davies, 2010; Wohlkonig et al , 2010; Domínguez-Gil et al , 2016). The C-type lysozymes use a dual catalytic acid-catalytic base mechanism.…”

Constructing and deconstructing the bacterial cell wall

“…283,284 Accordingly, we have undertaken extensive mechanistic and structural studies of this family. These efforts have led us to the discovery that an ABC-transporter-like regulatory module is present in the structure of the P. aeruginosa MltF LT 285 ; that the P. aeruginosa SltB1 LT has a catenane dimer structure; that the P. aeruginosa Slt LT 286 has the same compelling doughnut-shape as is seen in other Slt LTs whose catalytic activity is governed by important conformational changes 234,[287][288][289] ; and to a computational mechanism for the smallest LT of E. coli, MltE, 233 based on our crystallographic analyses. 290,291 The assertion of an intimate relationship between the LT family and β-lactam antibiotic efficacy has irrefutable support.…”