Secretory nucleoside diphosphate kinases from both intra- and extracellular pathogenic bacteria are functionally indistinguishable

Abstract

Institute of Microbial Technology, CSIR, Sector 39A, Chandigarh 160 036, IndiaNucleoside diphosphate kinase (NDK), responsible for the maintenance of NTP pools, is an ATP-utilizing enzyme secreted by different pathogens. We found that NDK from Salmonella enterica serovar Typhimurium (S. Typhimurium) is also secretory in nature. Secretory NDK is known to play a crucial role in the survival of pathogenic microbes within host cells through their interaction with extracellular ATP. To elucidate this aspect, we assessed the contribution of secretory products containing NDK from intracellular (Mycobacterium tuberculosis and S. Typhimurium) and extracellular (Vibrio cholerae) pathogens to the process of ATP-induced J774 mouse macrophage cell lysis by monitoring lactate dehydrogenase (LDH) release in the culture medium. Compared with an untreated control, our results demonstrate that S. Typhimurium secretory products caused a greater than twofold decrease in LDH release from J774 macrophage cells treated with ATP. Furthermore, the secretory products from an ndk-deleted strain of S. Typhimurium did not display such behaviour. Contrary to this observation, the secretory products containing NDK of V. cholerae were found to be cytotoxic to J774 cells. At the amino acid level, the sequences of both the NDKs (S. Typhimurium and V. cholerae) exhibited 65 % identity, and their biochemical characteristics (autophosphorylation and phosphotransfer activities) were indistinguishable. However, to our surprise, the secretory product of an ndk-deleted strain of S. Typhimurium, when complemented with V. cholerae ndk, was able to prevent ATP-induced cytolysis. Taken together, our results unambiguously imply that the intrinsic properties of secretory NDKs are identical in intra-and extracellular pathogens, irrespective of their mode of manifestation.

Nucleoside-Diphosphate-Kinase of P. gingivalis is Secreted from Epithelial Cells In the Absence of a Leader Sequence Through a Pannexin-1 Interactome

“…Currently it is recognized that NDKs are present in a vast number of species from bacteria to human and they can perform multifunctional duties within the organism1234. All presently sequenced and characterized NDKs represent tetra- or hexamers in their active enzyme form, with subunit size of 11–18 kDa234. Similar to other leaderless proteins that are shown to be secreted, there is still not much known on the specific pathways of NDK secretion, both on the microbial species level, as well as the higher organisms’ levels such as human1.…”

“…Nucleoside-diphosphate-kinases (NDKs) are a family of multifunctional enzymes that are evolutionarily highly conserved among numerous species from bacteria to humans12. One of their main functions is to hydrolyze nucleotide triphosphates (NTP), such as ATP, by catalyzing the transfer of orthophosphate residues, and are essential for proper DNA synthesis and housekeeping of cellular functions3.…”

“…One of their main functions is to hydrolyze nucleotide triphosphates (NTP), such as ATP, by catalyzing the transfer of orthophosphate residues, and are essential for proper DNA synthesis and housekeeping of cellular functions3. NDKs are small (~11–18 kDa), mainly cytoplasmic proteins, usually forming polymers (tetra- or hexamers) and having no known signal leader sequence, thus currently are unassigned to a specific secretion pathway234. The significance of NDK, including the human NDK species, encoded by the nm23 gene, has been suggested for the pathogenesis of several different forms of severe chronic diseases, including cancers567.…”

Role of extracellular nucleotides in the immune response against intracellular bacteria and protozoan parasites

“…Other intracellular bacteria, such as Mycobacterium tuberculosis and Porphyromonas gingivalis , also inhibit P2X7-mediated apoptosis of macrophages or primary gingival epithelial cells (GECs) apoptosis [45–47], which may be mediated by a homolog of nucleoside diphosphate kinase (Ndk) secreted by the bacteria [45,47]. It is possible that the P. gingivalis Ndk may also inhibits inflammation, since ATP treatment induces NLRP3-dependent caspase-1 activation and release of IL-1β in infected GECs cells [48].…”

Effect of Rocket (Eruca sativa) Extract on MRSA Growth and Proteome: Metabolic Adjustments in Plant-Based Media

“…Although NDK proteins have moderately similar sequence identities, they are highly conserved in prokaryotes and eukaryotes performing similar functions (Sundin et al, 1996). In bacteria, extracellular secretion of NDK has been reported in several pathogens such as Porphyromonas gingivalis (Yilmaz et al, 2008), Pseudomonas aeruginosa, Mycobacterium tuberculosis (Chakrabarty, 1998), and S. enterica serovar Typhimurium (Dar et al, 2011). It appears that apart from its major role in supporting growth and colonization of bacteria, NDK takes also part in other functions like virulence, cell signaling and polysaccharide synthesis, as demonstrated in Mycobacterium and Pseudomonas species (Chakrabarty, 1998).…”