Adhesion Proteins: Keeping Bacteria in Their Place
Many bacteria produce outer membrane-localized proteins that adhere them to varying biotic and abiotic substrates. Such interactions are crucial for the life-cycle of many microorganisms, promoting retention in high-nutrient locations and the formation of communities, called biofilms. From a human perspective, microbial adhesion and biofilm formation are often detrimental, causing chronic antibiotic-resistant infections, the corrosion and clogging of machinery, and the spoiling of resources. It is in the best interest of human health and industry to understand the molecular connections that allow these natural phenomena to take place, and how they can be augmented or sabotaged. As such, the purpose of this thesis was to interrogate the structure/function of several adhesion proteins produced by Gram-negative bacteria. Two of the three adhesion protein examples chosen are part of the Repeats-In-Toxin family, which relies on the type I secretion system for proper localization and cell-membrane retention. The examples include: 1) The 1.5-MDa ice-binding protein from the Antarctic bacterium Marinomonas primoryensis. The protein’s remarkable size can be attributed to one region that holds ~120 tandem repeat domains. Structure determination of a four-repeat segment confirmed these domains to be immunoglobulin-like β-sandwiches that bind calcium ions for both proper folding and rigidity, facilitating the region’s proposed role of extension. 2) The adhesion protein from the oil-eating bacterium Marinobacter hydrocarbonoclasticus, which houses a proposed PA14 domain close to its C-terminal tip. This domain was shown to bind sugar through a combination of X-ray crystallography and a custom-made competition assay. Potential applications for this domain as a dextran-affinity tag were also explored. And 3) an example from a different protein family was characterized due to the incorporation of a DUF3494 – a well-known ice-binding domain – into its distal tip. The ice-binding activity of this DUF3494 was confirmed and characterized, adding to the ongoing intrigue of this widespread and functionally-variable domain.