Accumulation of Poly(3-hydroxybutyrate) Helps Bacterial Cells to Survive Freezing.

Stanislav Obruca,Vladislav Krzyzanek,Ota Samek,Petr Sedlacek,Dan Kucera,Pavla Benesova,Kamila Hrubanova,Filip Mravec,Ivana Marova,Guo-Qiang Chen

doi:10.1371/journal.pone.0157778

Abstract

Accumulation of polyhydroxybutyrate (PHB) seems to be a common metabolic strategy adopted by many bacteria to cope with cold environments. This work aimed at evaluating and understanding the cryoprotective effect of PHB. At first a monomer of PHB, 3-hydroxybutyrate, was identified as a potent cryoprotectant capable of protecting model enzyme (lipase), yeast (Saccharomyces cerevisiae) and bacterial cells (Cupriavidus necator) against the adverse effects of freezing-thawing cycles. Further, the viability of the frozen–thawed PHB accumulating strain of C. necator was compared to that of the PHB non-accumulating mutant. The presence of PHB granules in cells was revealed to be a significant advantage during freezing. This might be attributed to the higher intracellular level of 3-hydroxybutyrate in PHB accumulating cells (due to the action of parallel PHB synthesis and degradation, the so-called PHB cycle), but the cryoprotective effect of PHB granules seems to be more complex. Since intracellular PHB granules retain highly flexible properties even at extremely low temperatures (observed by cryo-SEM), it can be expected that PHB granules protect cells against injury from extracellular ice. Finally, thermal analysis indicates that PHB-containing cells exhibit a higher rate of transmembrane water transport, which protects cells against the formation of intracellular ice which usually has fatal consequences.

Highlights

Microorganisms are exposed to a series of various stress factors in the environment, among which great temperature oscillations are very frequent
In our previous study we identified 3HB as a very potent chemical chaperone capable of protecting lipase and lysozyme against adverse effects of heat and oxidative damage [16]
Cryoprotective ability has been observed for a wide spectrum of compatible solutes which are produced and accumulated by various microorganisms to cope with ubiquitous environmental stresses such as high osmolality or temperature fluctuations [29, 30]

Summary

Introduction

Microorganisms are exposed to a series of various stress factors in the environment, among which great temperature oscillations are very frequent. It should be noted that approximately 80% of our planet’s biosphere is permanently cold with average temperatures below 5°C and that even in the remaining regions the temperature fluctuates wildly, occasionally falling close to, or even below 0°C [1]. Many microorganisms have developed sophisticated strategies to help them endure low temperatures. These include the capacity to produce and accumulate cryoprotectants, substances which are able to protect cells from the adverse effects of freezing and low temperature [2]. There are numerous low molecular weight solutes (e.g. amino acids and their derivatives, sugars, ectoines and their derivatives, etc.) as well as high.

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