asymmetric unit</scene> ((6OYH))' size='340' side='right' caption='Crystal structure of two MraY dimers'>

Introduction

MraY

MraY, called also phospho-N-acetylmuramoyl-pentapeptide-transferase or UDP-MurNAc-pentapeptide phosphotransferase, with EC number 2.7.8.13, is an integral membrane enzyme involved in peptidoglycan biosynthesis ^{[1] [2]}.MraY is encoded by the mraY gene and belongs to a subfamily of the polyprenyl-phosphate N-acetyl hexosamine 1-phosphate transferase (PNPT) superfamily ^[3]. MRAY is a promising candidate for the development of new antibiotics. In fact, it is the target of five classes of natural nucleoside inhibitors with potent antibacterial activity: the liposidomycins/caprazamycins, capuramycins, mureidomycins, muraymycins, and tunicamycins ^{[2] [4]}.The structure presented in this page correspond to the MraY protein from the thermophile Aquifex aeolicus strain VF5 (MraYAA) in complex with carbacaprazamycin.MraYAA was expressed in Escherichia coli.

Carbacaprazamycin

is a chemically stable analog of caprazamycin nucleoside inhibitors ^[5]. One particular characteristic of caprazamycin nucleoside inhibitors is that they have uridine on their structure. It has been shown that Mray inhibitors contains a uridine moiety ^[6]

Function

MraY is a critical enzyme in peptidoglycan biosynthesis. Peptidoglycan is an essential component of the cell wall of Gramnegative and Gram-positive bacteria ^[7]. The cell wall provides bacteria a structural support and protection. In particular, it allows bacteria to maintain their cell shape at different osmotic pressures ^[8]. Peptidoglycan is a cross-linked polymer of carbohydrates an amino acids and due to its biological relevance in Bacteria, it has been a major target for antibiotics ^{[3] [7]}.

Peptidoglycan biosynthesis involves three main stages. MraY is responsible for the second stage. First, the peptidoglycan precursor UDP-Nacetylmuramoyl (MurNAc)–pentapeptide is synthesized in the cytosol. Second, this hydrophilic precursor is attached to a lipid carrier, and the complex lipid carrier-precursor is transported, through the membrane, to the periplasm. Third, the peptidoglycan precursors are polymerized to form the cell wall. MraY catalyzes the transfer of phospho-MurNAc-pentapeptide from hydrophilic substrate UDP-MurNAc-pentapeptide to the lipid carier (C55-P) in the presence of a Mg2+ cofactor. The product is the undecaprenyl-pyrophosphoryl-MurNAcpentapeptide, also known as lipid I ^{[3] [6] [9]}.

Structure

Asymmetric and biological unit

The basal 3D scene shown on this page corresponds to the , which contains 4 biological units of MraYAA (colors), which each of them is fused with the NB7 nanobody (teal). Mashalidis EH, Kaeser Bet al. , who solved this 3D structure, were screening antibodies. They determine that the presence of NB7 does not affect MraYAA enzymatic activity or inhibition by carbacaprazamycin ^[6].

MraY-Carbacaprazamycin structure

In bacteria, MraYAA is in the form of a dimer, which actually what researchers found in the first 3D structure of MraYAA ^[3]. Its N and the C termini are located on the periplasmic side. This protein is made of four extracellular loops, five cytoplasmic loops (named A,B, C, D and E) and ten transmembrane helices named TM1 to TM10. Though, TM9 is cleaved by a glycin residue into two helical segments named TM9a and TM9b. Furthermore, there is an additional helix between TM9b and TM10, which is only 11 residues long and is called TM9c. TM9c contains a (H324,325 and 326) which plays a role in the enzyme’s substrat selectivity and is a part of the catalytic site ^[1]. They intereact with carbacaprazamycin, and also with other inhibitors such as muraymycin D2 (MD2) . This small loop binds two Ni2+ ions, one on the two first histidines and the other one on the last one. Another part of this site is the residues corresponding to ^[4].TM5–TM10 and loops C and D also play a role in the catalytic site and contain many polar and charged amino acids residues ^[1]. Also, some polar and charged amino acids on TM9b and loop E are pointing toward the active site, making it even more hydrophilic.

is made of uridine, 5-aminoribosyl, diazepanone, and aliphatic tail moieties. The diazepanone ring system makes relatively few interactions with the protein. Carbacapzazamycin contains an uridine binding pocket which is formed by amino acid residues in Loop C ( ). This pocket is capped off by a π–π stacking interaction with in Loop D. forms an additional hydrogen bond with the uracil moiety. Next to the uridine binding site, there is a second binding pocket lined with amino acid residues, called the uridine-adjacent pocket. The 5-aminoribose moiety of carbacaprazamycin forms an extensive hydrogen bond network in the uridine-adjacent pocket. from those results we can observe that the uracil moiety binds to an enclosed pocket on the cytoplasmic face of MraY, while the ribosyl moiety is more solvent exposed ^{[5] [6]}.

Relevance

Drug resistant bacteria are the cause of death of millions of people worldwide. In the USA alone, hospital infections associated with antibiotic-resistant pathogens cause 99 000 deaths per year^[10]. The development of new antibiotics with new mechanism of action is urgent. This crystal structure, 6OYH, serve as a generalizable MraY structural model to study and better understand the action of the nucleosides inhibitors. Moreover, the sttructural analysis of the binding of carbacaprazamycin to MraYAA provides a better understanding of the chemical logic of MraY inhibition, which can help in the development of novel approaches for the design of antibiotics targeting MraY.

3D related structures

• 4j72:Crystal Structure of polyprenyl-phosphate N-acetyl hexosamine 1-phosphate transferase (MraY)
• 6oyz:Crystal structure of MraY bound to capuramycin
• 5ckr:Crystal Structure of MraY in complex with Muraymycin D2
• 6oz6:Crystal structure of MraY bound to 3'-hydroxymureidomycin A
• 5jnq:MraY tunicamycin complex

This is a sample scene created with SAT to by Group, and another to make of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.