Abstract
BackgroundThiamine triphosphate (ThTP) exists in most organisms and might play a role in cellular stress responses. In E. coli, ThTP is accumulated in response to amino acid starvation but the mechanism of its synthesis is still a matter of controversy. It has been suggested that ThTP is synthesized by an ATP-dependent specific thiamine diphosphate kinase. However, it is also known that vertebrate adenylate kinase 1 catalyzes ThTP synthesis at a very low rate and it has been postulated that this enzyme is responsible for ThTP synthesis in vivo.ResultsHere we show that bacterial, as vertebrate adenylate kinases are able to catalyze ThTP synthesis, but at a rate more than 106-fold lower than ATP synthesis. This activity is too low to explain the high rate of ThTP accumulation observed in E. coli during amino acid starvation. Moreover, bacteria from the heat-sensitive CV2 strain accumulate high amounts of ThTP (>50% of total thiamine) at 37°C despite complete inactivation of adenylate kinase and a subsequent drop in cellular ATP.ConclusionThese results clearly demonstrate that adenylate kinase is not responsible for ThTP synthesis in vivo. Furthermore, they show that E. coli accumulate large amounts of ThTP under severe energy stress when ATP levels are very low, an observation not in favor of an ATP-dependent mechanisms for ThTP synthesis.
Highlights
Thiamine triphosphate (ThTP) exists in most organisms and might play a role in cellular stress responses
On the other hand, when the bacteria were incubated in minimal medium devoid of any carbon source, we noticed the appearance of a new compound that was identified as adenosine thiamine triphosphate (AThTP) [9]
As AK is a possible candidate for catalyzing ThTP synthesis, it is important to determine whether bacterial AKs are able to catalyze this reaction and, if they do, to know whether the specific activity of, in particular, E. coli AK is sufficient to account for the relatively high rate of ThTP production measured in vivo in this organism
Summary
Thiamine triphosphate (ThTP) exists in most organisms and might play a role in cellular stress responses. Overexpression in E. coli of a specific soluble mammalian thiamine triphosphatase (ThTPase), that we previously characterized [6,7,8], prevented ThTP accumulation and induced the appearance of an intermediate plateau in bacterial growth [5]. This suggested that ThTP may be required for the rapid adaptation of bacteria to amino acid starvation. ThTP and AThTP never accumulate simultaneously in high amounts, suggesting that the two compounds may act as specific alarmones, responding to different conditions of cellular stress
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