Salmonella enterica serovar Typhimurium has three transketolase enzymes contributing to the pentose phosphate pathway

Abstract

The genus is responsible for many illnesses in humans and other vertebrate animals. We report here that serovar Typhimurium harbors three transketolases that support the non-oxidative branch of the pentose phosphate pathway. BLAST analysis identified two genes, _ and _, that together encode a putative transketolase (TktC) with 46-47% similarity to the known TktA and TktB isoforms. Assessing the mRNA and protein expression for each of the three transketolases, we determined that all are expressed in WT cells and regulated to varying extents by the alternative sigma factor RpoS. Enzyme assays with lysates from WT and transketolase-knockout strains established that TktA is responsible for >88% of the transketolase activity in WT cells. We purified recombinant forms of each isoenzyme to assess the kinetics for canonical transketolase reactions. TktA and TktB had comparable values for (539-1362 μm NADH consumed/s), (80-739 μm), and catalytic efficiency (1.02 × 10-1.06 × 10 m/s) for each substrate tested. The recombinant form of TktC had lower values (23-120 μm), whereas the (7.8-16 μm NADH consumed/s) and catalytic efficiency (5.58 × 10 to 6.07 × 10 m/s) were 10-100-fold lower. Using a murine model of infection, we showed that a strain lacking all three transketolases is avirulent in C57BL/6 mice. These data provide evidence that Typhimurium possesses three transketolases that contribute to pathogenesis.

CrrB Positively Regulates High-Level Polymyxin Resistance and Virulence in Klebsiella pneumoniae

“…The role of these pathways in virulence of Gram-negative organisms is not completely clear, but work has shown that the PPP is activated during infection. For example, in Salmonella enterica, the KO of the transketolase enzymes within the PPP renders the resulting mutant avirulent in a mouse model (Shaw et al, 2018;Spaans et al, 2015). One potential explanation for this could be that the upregulation of these pathways shunts carbon intermediates toward LPS biosynthesis, a key virulence factor in Gram-negative bacteria (Stincone et al, 2015).…”

Prevalence of Multidrug-Resistant Salmonella enterica Serovars in Buffalo Meat in Egypt

“…At present, Salmonella enterica serovars are reported worldwide as the most common cause of human gastroenteritis associated with the consumption of contaminated foods. S. enterica is a very important food-borne bacteria that results in several public health troubles for humans and animals [ 7 ]. It is considered to be the second bacterial foodborne disease infecting humans.…”

Transketolase is involved in the control of Sigma B during chronic infection byStaphylococcus aureus

“…Samples were centrifuged at 12,000 g and 4 °C for 10 min to remove cell debris and protein concentration was determined using BCA assay with BSA as a standard (Thermo Fisher Scientific). TKT activity was analysed as described previously (76). Briefly, each enzymatic reactions were performed at a final volume of 100 µl containing 20 µl of supernatant, 50 mM Tris (pH 7.5), 0.24 mM MgCl 2 , 10 µM thiamine pyrophosphate, 0.5 mM NADH, 3 units of glycerol-3-phosphate dehydrogenase, 10 units of triosephosphate isomerase, 0.5mM D-xylulose-5-phosphate, and 0.5mM D-erythrose-4-phosphate (all reagents obtained from Sigma-Aldrich).…”