Molybdate pumping into the molybdenum storage protein via an ATP-powered piercing mechanism

The molybdenum storage protein (MoSto) deposits large amounts of molybdenum as polyoxomolybdate clusters in a heterohexameric (αβ)3cage-like protein complex under ATP consumption. Here, we suggest a unique mechanism for the ATP-powered molybdate pumping process based on X-ray crystallography, cryoelectron microscopy, hydrogen-deuterium exchange mass spectrometry, and mutational studies of MoSto fromAzotobacter vinelandii. First, we show that molybdate, ATP, and Mg2+consecutively bind into the open ATP-binding groove of the β-subunit, which thereafter becomes tightly locked by fixing the previously disordered N-terminal arm of the α-subunit over the β-ATP. Next, we propose a nucleophilic attack of molybdate onto the γ-phosphate of β-ATP, analogous to the similar reaction of the structurally related UMP kinase. The formed instable phosphoric-molybdic anhydride becomes immediately hydrolyzed and, according to the current data, the released and accelerated molybdate is pressed through the cage wall, presumably by turning aside the Metβ149 side chain. A structural comparison between MoSto and UMP kinase provides valuable insight into how an enzyme is converted into a molecular machine during evolution. The postulated direct conversion of chemical energy into kinetic energy via an activating molybdate kinase and an exothermic pyrophosphatase reaction to overcome a proteinous barrier represents a novelty in ATP-fueled biochemistry, because normally, ATP hydrolysis initiates large-scale conformational changes to drive a distant process.

UMP Kinase Regulates Chloroplast Development and Cold Response in Rice

Pyrimidine nucleotides are important metabolites that are building blocks of nucleic acids, which participate in various aspects of plant development. Only a few genes involved in pyrimidine metabolism have been identified in rice and the majority of their functions remain unclear. In this study, we used a map-based cloning strategy to isolate a UMPK gene in rice, encoding the UMP kinase that phosphorylates UMP to form UDP, from a recessive mutant with pale-green leaves. In the mutant, UDP content always decreased, while UTP content fluctuated with the development of leaves. Mutation of UMPK reduced chlorophyll contents and decreased photosynthetic capacity. In the mutant, transcription of plastid-encoded RNA polymerase-dependent genes, including psaA, psbB, psbC and petB, was significantly reduced, whereas transcription of nuclear-encoded RNA polymerase-dependent genes, including rpoA, rpoB, rpoC1, and rpl23, was elevated. The expression of UMPK was significantly induced by various stresses, including cold, heat, and drought. Increased sensitivity to cold stress was observed in the mutant, based on the survival rate and malondialdehyde content. High accumulation of hydrogen peroxide was found in the mutant, which was enhanced by cold treatment. Our results indicate that the UMP kinase gene plays important roles in regulating chloroplast development and stress response in rice.

Aliivibrio finisterrensis sp. nov., isolated from Manila clam, Ruditapes philippinarum and emended description of the genus Aliivibrio

Four strains isolated from cultured Manila clam, Ruditapes philippinarum, in the north-western coast of Spain were characterized phenotypically and genotypically. Phylogenetic analyses based on the 16S rRNA gene sequences indicated that these bacteria were closely related to Aliivibrio wodanis, Aliivibrio salmonicida, Aliivibrio fischeri and Aliivibrio logei with sequence similarities between 98.1 and 96.0 %. Phylogenetic analysis based on the RNA polymerase alpha chain (rpoA), RecA protein (recA), the α-subunit of bacterial ATP synthase (atpA) and the uridine monophosphate (UMP) kinase (pyrH) genes and fluorescent amplified fragment length polymorphism experiments clearly showed that these novel isolates form a tight genomic group different from any currently known Aliivibrio species. On the basis of phylogenetic analysis and phenotypic data, the four strains represent a novel taxon, for which the name Aliivibrio finisterrensis sp. nov. is proposed. Several phenotypic features were revealed that discriminate A. finisterrensis from other Aliivibrio species. The type strain is CMJ 11.1T (=CECT 7228T=LMG 23869T).