NDPK

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Introductions

The Nucleoside-diphosphate kinases (NDPK) gene is highly conserved in evolution, it presents complex and diverse biological functions. In addition to catalyzing the transfer of high-energy phosphate groups between adenosine triphosphate (NTP) and nucleoside diphosphate (NDP), NDPK also possesses NDPK and protein phosphotransferase activities and is involved in transcriptional regulation and signal transduction. The cDNA sequence of silkworm nucleoside diphosphate kinase was obtained from the silkworm pupae cDNA library. The cDNA sequence was 575 bp long and contained a 465 bp open reading frame sequence, encoding a nucleoside diphosphate kinase with 154 amino acid residues. The recombinant plasmid was constructed by inserting the cloned silkworm nucleoside diphosphate kinase gene into pET-28a, and the recombinant protein was induced to be expressed after transformation into Escherichia coli and purified by nickel ion affinity chromatography column to obtain recombinant silkworm nucleoside diphosphate kinase.

Functions

Nucleoside diphosphate kinases (NDPK) are nucleotide metabolizing enzymes encoded by the NME (also known as NM23) gene. NDPK is the ubiquitous enzyme that catalyzes the transfer of phosphate from nucleoside triphosphate (NTP) to nucleoside diphosphate (NDP), specifically GDP, through a ping-pong mechanism involving the formation of a phosphate-histidine intermediate. In this reaction, the primary donor of phosphate is ATP, which is supplied primarily by mitochondrial oxidative phosphorylation as its intracellular concentration is much higher than any other nucleoside triphosphate. In mammals, 10 genes have been identified as belonging to the NME/NDPK family. They encode proteins with one or two NDPK structural domains or a truncated NDPK structural domain that is ultimately associated with other structural domains of largely unknown function.

NDPK in Metastasis

Metastasis is the leading cause of death in cancer patients. The first step in metastasis is the transformation from carcinoma in situ to invasive carcinoma as the basement membrane is disrupted and tumor cells migrate infiltratively through the mesenchymal fiber type I collagen. The tumor cells then enter the circulation and exit to colonize secondary organs and form metastases. Although type 1 matrix metalloproteinase (MT1-MMP) is involved in this process, the mechanism of transformation from in situ to invasive carcinoma remains largely unknown. The identification of NME1 as the first metastasis suppressor gene26 has opened the field of this new class of metastasis dissemination regulators. Unlike tumor suppressor genes, metastasis suppressors inhibit in vivo metastasis without globally inhibiting primary tumor growth.

NDPK as a Protein Histidine Kinase

In addition to its recognized role as an NDP kinase, there is new evidence that NME/NDPK proteins act as proteohistidine kinases in mammals. For NME2/NDPK-B, at least two substrates were identified and studied in more detail: the β-subunit of the heterotrimeric G protein and the potassium channel KCa3.1. In the first case, NME2 forms a complex with β/γ to form a dimer of the heterotrimeric G protein in which a phosphate relay occurs from His118 of NME2 to His266 of the G β-subunit. This eventually leads to the formation of GTP from GDP, which results in the non-dependent activation of the receptor for G proteins. A second well-documented example is the NME2-mediated activation of the K⁺ channel KCa3.1, which induces cytokine production by CD⁴⁺ cells.

Conclusions

Over the past three decades, extensive analysis of the NDPK family has begun to reveal its multifaceted role in cytopathophysiology and underlying molecular mechanisms. Starting from simple kinase activities, the field is now faced with the complexity of 10 different isoforms, all with specific characteristics and often with alternative, additional activities. The role of these different isoforms undoubtedly requires a specific target and intracellular location. Most studies have addressed the specific roles of the two most abundantly expressed isoforms, NME1 and NME2, with particular emphasis on the anti-metastatic function of NME1.