9.A.74.  The Small Oligomeric Nitrate/Nitrite Uptake Transporter (NrtS) Family 

A putative silent gene of the freshwater cyanobacterium Synechococcus elongatus strain PCC 7942, encoding a small protein with two TMSs, was named nrtS, since its overexpression from an inducible promoter conferred nitrate uptake activity on the nitrate transport-less NA4 mutant of S. elongatus (Maeda et al. 2019). Homologs of NrtS, of 67-118 aas, are present in limited numbers of eubacteria including cyanobacteria and proteobacteria, but also actinobacteria of the Mycobacterium tuberculosis complex. When expressed in NA4, the NrtS homolog of the gamma-proteobacterium Marinomonas mediterranea took up nitrate with higher affinity than the S. elongatus NrtS (K_m of 0.49 mM vs. 2.5 mM). Among the 61 bacterial species carrying an nrtS gene, the marine cyanobacterium Synechococcus sp. strain PCC 7002 is unique in having two nrtS genes (nrtS1 and nrtS2) located in tandem on the chromosome. Coexpression of the two genes in NA4 resulted in nitrate uptake with a K_m(NO₃^-) of 0.15 mM, while expression of either of the two resulted in low-affinity nitrate uptake activity with K_m values of > 3 mM, indicating that NrtS1 and NrtS2 form a heteromeric transporter complex. The heteromeric transporter was shown to transport nitrite as well as nitrate. A Synechococcus sp. strain PCC 7002 mutant defective in the nitrate transporter (NrtP) showed a residual activity of nitrate uptake, which was ascribed to the NrtS proteins. Blue-native PAGE and immunoblotting analysis suggested a hexameric structure for the NrtS proteins (Maeda et al. 2019).

It is not known if these transporters function by a channel- or carrier-type mechanism.  On the one hand, no system with subunits of less than 3 TMSs has previously been shown to be a carrier although many are channels, but on the other hand, nitrate and nitrite are negatively charged, and therefore would not be easily taken up into the cytoplasm (membrane potential negative inside) if a channel-type mechanism were operative.  Further experimentation will be required to distinguish these possibilities. 

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