Robert Turner

The University of Sheffield | Sheffield · Department of Molecular Biology and Biotechnology

Understanding how bacteria grow and divide requires insight into both the molecular-level dynamics of ultrastructure and the chemistry of the constituent components. Atomic force microscopy (AFM) can provide near molecular resolution images of biological systems but typically provides limited chemical information. Conversely, while super-resolution...

Biopolymer composite cell walls maintain cell shape and resist forces in plants, fungi and bacteria. Peptidoglycan, a crucial antibiotic target and immunomodulator, performs this role in bacteria. The textbook structural model of peptidoglycan is a highly ordered, crystalline material. Here we use atomic force microscopy (AFM) to image individual g...

The bacterial cytoplasmic membrane is the interface between the cell and its environment, with multiple membrane proteins serving its many functions. However, how these proteins are organised to permit optimal physiological processes is largely unknown. Based on our initial findings that 2 phospholipid biosynthetic enzymes (PlsY and CdsA) localise...

The bacterial cell wall is essential for viability, but despite its ability to withstand internal turgor must remain dynamic to permit growth and division. Peptidoglycan is the major cell wall structural polymer, whose synthesis requires multiple interacting components. The human pathogenStaphylococcus aureusis a prolate spheroid that divides in th...

Antimicrobial resistance is recognized as one of the principal threats to public health worldwide, yet the problem is increasing. Methicillin–resistant Staphylococcus aureus (MRSA) are among the most difficult to treat in clinical settings due to the resistance to nearly all available antibiotics. The cyclic anionic lipopeptide antibiotic Daptomyci...

Atomic force microscopy (AFM) has been used extensively to characterize the surface structure and mechanical properties of bacterial cells. Extraction of the cell wall peptidoglycan sacculus enables AFM analysis exclusively of peptidoglycan architecture and mechanical properties, unobscured by other cell wall components. This has led to discoveries...

Unlabelled: Most bacterial cells are enclosed in a single macromolecule of the cell wall polymer, peptidoglycan, which is required for shape determination and maintenance of viability, while peptidoglycan biosynthesis is an important antibiotic target. It is hypothesized that cellular enlargement requires regional expansion of the cell wall throug...

The polyphenol (-)-epicatechin gallate (ECg) inserts into the cytoplasmic membrane (CM) of methicillin-resistant Staphylococcus aureus (MRSA) and reversibly abrogates resistance to β-lactam antibiotics. ECg elicits an increase in MRSA cell size and induces thickened cell walls. As ECg partially delocalizes penicillin-binding protein PBP2 from the s...

The nanoscale mechanical properties of live Staphylococcus aureus cells during different phases of growth were studied by atomic force microscopy. Indentation to different depths provided access to both local cell wall mechanical properties and whole-cell properties, including a component related to cell turgor pressure. Local cell wall properties...

Bacterial cell division is a fundamental process that requires the coordinated actions of a number of proteins which form a complex macromolecular machine known as the divisome. The membrane-spanning proteins DivIB and its orthologue FtsQ are essential divisome components in Gram-positive and Gram-negative bacteria, respectively. However, the role...

Human skin fatty acids are a potent aspect of our innate defenses, giving surface protection against potentially invasive organisms. They provide an important parameter in determining the ecology of the skin microflora, and alterations can lead to increased colonization by pathogens such as Staphylococcus aureus. Harnessing skin fatty acids may als...

Peptidoglycan performs the essential role of resisting turgor in the cell walls of most bacteria. It determines cell shape, and its biosynthesis is the target for many important antibiotics. The fundamental chemical building blocks of peptidoglycan are conserved: repeating disaccharides cross linked by peptides. However, these blocks come in many v...

Cellular integrity and morphology of most bacteria is maintained by cell wall peptidoglycan, the target of antibiotics essential in modern healthcare. It consists of glycan strands, cross-linked by peptides, whose arrangement determines cell shape, prevents lysis due to turgor pressure and yet remains dynamic to allow insertion of new material, and...

Sacculus immobilised on Cell-Tak/PDMS and stretched showing maintenance of features under strain

In situ Atl amidase digest of sacculi under buffer A showing loss of feature orientation and punctation of material (10 nm height scale).

Supplementary Figures S1-S3 and Supplementary Table S1

In situ lysozyme digest of sacculi under buffer A showing loss of feature orientation and enlargement of pores (10 nm height scale).

Division in Staphylococci occurs equatorially and on specific sequentially orthogonal planes in three dimensions, resulting, after incomplete cell separation, in the 'bunch of grapes' cluster organization that defines the genus. The shape of Staphylococci is principally maintained by peptidoglycan. In this study, we use Atomic Force Microscopy (AFM...

Preparation of vital bacteria for atomic force microscope study under aqueous fluid, such as physiological buffer or bacterial growth medium, presents challenges as cells will often desorb from the supporting surface or be dislodged by the atomic force microscope tip during imaging. An established method of immobilizing coccoid bacteria is to trap...

Monitoring higher harmonics of the drive frequency in amplitude-modulation atomic force microscopy can give extra information on local surface properties. The first to fourth harmonics inclusive were monitored on the surface of individual Staphylococcus aureus bacteria and on the polycarbonate filter in which they were trapped. The second harmonic...