Molecular adaptation and salt stress response of Halobacterium salinarum cells revealed by neutron spectroscopy

Abstract

Halobacterium salinarum is an extreme halophile archaeon with an absolute requirement for a multimolar salt environment. It accumulates molar concentrations of KCl in the cytosol to counterbalance the external osmotic pressure imposed by the molar NaCl. As a consequence, cytosolic proteins are permanently exposed to low water activity and highly ionic conditions. In non-adapted systems, such conditions would promote protein aggregation, precipitation, and denaturation. In contrast, in vitro studies showed that proteins from extreme halophilic cells are themselves obligate halophiles. In this paper, adaptation via dynamics to low-salt stress in H. salinarum cells was measured by neutron scattering experiments coupled with microbiological characterization. The molecular dynamic properties of a proteome represent a good indicator for environmental adaptation and the neutron/microbiology approach has been shown to be well tailored to characterize these modifications. In their natural setting, halophilic organisms often have to face important variations in environmental salt concentration. The results showed deleterious effects already occur in the H. salinarum proteome, even when the external salt concentration is still relatively high, suggesting the onset of survival mechanisms quite early when the environmental salt concentration decreases.

Surviving salt fluctuations: stress and recovery in Halobacterium salinarum, an extreme halophilic Archaeon

“…As discussed previously, the molecular dynamic state of the cellular proteome represents a robust indicator of cellular fitness 14,15,31 . Macromolecular denaturation through unfolding leads to lower effective force constants indicating less rigid structures.…”

“…In the case of Hbt. salinarum we have shown that below 2 M NaCl (a value which induces unfolding of the model Haloarcula marismortui enzyme, MalDH, in absence of cofactors) halophilic cells suffer a high mortality rate 15 . In the same study, molecular dynamics parameters of the proteome were measured in vivo by neutron scattering for cultures exposed to various lower salt concentrations.…”

“…marismortui and of the native proteome from Hbt. salinarum [12][13][14][15][16] indicated that halophilic proteins display specific biophysical adaptation properties to high salt reflected by higher structural resilience, compared to mesophilic systems. It is likely, therefore, that most halophilic proteins display an obligatory requirement for high salt conditions in order to be active and stable.…”

High protein flexibility and reduced hydration water dynamics are key pressure adaptive strategies in prokaryotes

“…Consequently, only few of them were studied so far by means of incoherent neutron scattering, among them Escherichia coli 78 ( E. coli ), Haloarcula marismortui 9, red blood cells10, Halobacterium salinarum 1112 and very recently Shevanella Oneidensis 13. The advantages of neutron scattering consist in the huge incoherent cross section of hydrogen with respect to other atoms present in biological samples, which allows studying cellular constituents8 as well as water dynamics14, and in the fact that unlike X-rays, thermal neutrons have such a low energy that they do not induce radiation damage to the sample.…”

Regulação pós-transcricional em Halobacterium salinarum NRC-1: caracterização do gene VNG_RS05855

“…Em bactérias, a expressão diferencial de seus membros é associado com mecanismos de regulação pós-transcricionais(Owolabi e Rosen, 1990).Baseados nos estudos feitos por Beer e col, onde foram examinadas variações na composição do meio CM padrão do crescimento de H. salinarum NRC-1(Beer, Wurtmann e Baliga, 2014), testamos duas composições distintas: 86% de sais em relação ao meio CM (meio A), e 72% (meio B). A diminuição de sais no meio de crescimento de H. salinarum NRC-1 promovem mudanças nas taxas de crescimento, na morfologia da célula, conteúdo de vesículas de gás e regulação de genes vinculados ao metabolismo anaeróbico(Beer, Wurtmann e Baliga, 2014;Vauclare et al, 2015).No meio A, observamos que a partir de 50 horas de crescimento a linhagem nocaute apresenta menor OD 600nm do que a linhagem controle (Figura 4.8). Isto pode indicar uma maior sensibilidade a variações na concentração de sal na transição entre as fases exponencial e estacionária do crescimento, onde o metabolismo de H. salinarum NRC-1 pode mudar de Porém, só se detectam pequenas diferenças entre as linhagens a partir das 91 horas (Figura 4.8).…”