Evidence that TP_0144 of Treponema pallidum Is a Thiamine-Binding Protein

Abstract

Thiamine pyrophosphate (TPP), the biologically active form of thiamine (also known as vitamin B1), is an essential cofactor for several important enzymes involved in carbohydrate metabolism, and therefore, it is required for all living organisms. We recently found that a thiamine-binding protein (TDE_0143) is essential for the survival ofTreponema denticola, an important bacterial pathogen that is associated with human periodontitis. In this report, we provide experimental evidence showing that TP_0144, a homolog of TDE_0143 from the syphilis spirocheteTreponema pallidum, is a thiamine-binding protein that has biochemical features and functions that are similar to those of TDE_0143. First, structural modeling analysis reveal that both TDE_0143 and TP_0144 contain a conserved TPP-binding site and share similar structures to the thiamine-binding protein ofEscherichia coli. Second, biochemical analysis shows that these two proteins bind to TPP with similar dissociation constant (Kd) values (TDE_0143,Kdof 36.50 nM; TP_0144,Kdof 32.62 nM). Finally, heterologous expression of TP_0144 in a ΔTDE_0143 strain, a previously constructedTDE_0143mutant ofT. denticola, fully restores its growth and TPP uptake when exogenous thiamine is limited. Collectively, these results indicate that TP_0144 is a thiamine-binding protein that is indispensable forT. pallidumto acquire exogenous thiamine, a key nutrient for bacterial survival. In addition, the studies shown in this report further underscore the feasibility of usingT. denticolaas a platform to study the biology and pathogenicity ofT. pallidumand probably other uncultivable treponemal species as well.