Magnetic resonance tells microbiology where to go; bacterial teichoic acid protects liquid water at sub-zero temperatures

Abstract

Numerous chemical additives lower the freezing point of water, but life at sub-zero temperatures is sustained by a limited number of biological cryoprotectants. Antifreeze proteins in fish, plants, and insects provide protection to a few degrees below freezing. Microbes have been found to survive at even lower temperatures, although, with a few exceptions, antifreeze proteins are missing. Survival has been attributed to external factors, such as high salt concentration (brine veins) and adhesion to particulates or ice crystal defects. Teichoic acid is a phosphodiester polymer ubiquitous in Gram positive bacteria, composing 50% of the mass of the bacterial cell wall and excreted into the extracellular space of biofilm communities. We have found that when bound to the peptidoglycan cell wall (wall teichoic acid) or as a free molecule (lipoteichoic acid), teichoic acid is surrounded by liquid water at temperatures significantly below freezing. Using solid-state NMR, we are unable to collect 31 P CPMAS spectra for frozen solutions of lipoteichoic acid at temperatures above -60 °C. For wall teichoic acid in D 2 O, signals are not seen above -30 °C. These results can be explained by the presence of liquid water, which permits rapid molecular motion to remove 1 H/ 31 P dipolar coupling. 2 H quadrupole echo NMR spectroscopy reveals that both liquid and solid water are present. We suggest that teichoic acids could provide a shell of liquid water around biofilms and planktonic bacteria, removing the need for brine veins to prevent bacterial freezing. IntroductionNature relies on chemistry to maintain liquid water for life at subfreezing temperatures. The current paradigms use thermodynamic and/or kinetic arguments for cryosurvival in mammals, plants and insects.(1-3) At the onset of freezing, frogs excrete large amounts of glucose to lower the freezing point of water while preventing the formation of large ice crystals. Fish, plants, and insects inhibit ice formation with antifreeze proteins (AFP) that bind to the growth face of ice crystals. Enthalpic contributions are accompanied by entropic factors from a disorganized chemical system.(4) These factors slow the kinetics of crystallization such that AFPs perform their task at very low concentration, characterized as a non-colligative depression of the freezing point.Little is known of the cryoprotection chemistry for the predominate organisms in cold environments: microbes.(5, 6) Bacteria are found in nearly all low temperature environments and must prevent ice formation to ensure survival. Microbial communities have been discovered in ice,(7) snow,(8) permafrost,(9) and at the polar ice caps.(10, 11) Analysis of core samples show the presence of viable life hundreds of feet below the ice surface. Samples of frozen tundra have yielded various Gram positive, Gram negative, and Archaea species. (9, 12, 13) Here, Gram-negative Cryseobacterium and Gram-positive Enterococci were found to have a symbiotic interaction. The basis for cryosurvival is unknown, but macromolecules an...

RNA World meets Snowball Earth

“…27 It was during low-temperature NMR studies with D 2 O solutions of TA/MgCl 2 that we made the serendipitous discovery that TA protects liquid water at temperatures well below the freezing point. 7 Control experiments related to this work revealed that RNA also protects liquid water at -20 °C whereas DNA does not. These data are described below and used to design the experimental methods.…”

“…We are well-prepared to perform this work after making the serendipitous antifreeze discovery of RNA and TA. 7 Antifreeze proteins (AFPs) are not found in microbes thus finding antifreeze properties for RNA (found in all life) and bacterial TA (found in Gram+ bacteria) fills a void in cryobiology. Nevertheless, arctic fish have AFPs found in two distinct chemical categories.…”

Thermophiles and Psychrophiles in Nanotechnology

“…Eurypsychrophilic methanogens have also been found to possess temperature-dependent transcription factors that respond to variations in temperature by changing proteome composition (Goodchild et al 2004). Finally, extracellular polysaccharide substances (EPS) and teichoic acid are cryoprotectants, and intracellular fumarate and glycerol are chaotropes commonly produced upon cold shock (Rice et al 2008;Marx et al 2009;Chin et al 2010).…”