Two stages of substrate discrimination dictate selectivity in the MetNI-Q ABC transporter system.

The MetNI-Q importer, an ATP-binding cassette (ABC) transporter, mediates the uptake of both L- and D- enantiomers of methionine. Original uptake studies show a strong preference for L-Met over D-Met, but the molecular basis of this selectivity is unclear. In this work, we systematically examine substrate discrimination by the MetNI transporter and MetQ substrate binding protein using an array of biophysical and biochemical techniques.

Titratable transmembrane residues and a hydrophobic plug are essential for manganese import via the Bacillus anthracis ABC transporter MntBC-A.

All extant life forms require trace transition metals (e.g., Fe, Cu, and Mn) to survive.

Substrate recognition and ATPase activity of the cysteine/cystine ABC transporter YecSC-FliY.

Sulfur is essential for biological processes such as amino acid biogenesis, iron-sulfur cluster formation, and redox homeostasis. To acquire sulfur-containing compounds from the environment, bacteria have evolved high-affinity uptake systems, predominant among which is the ABC transporter family. Theses membrane-embedded enzymes use the energy of ATP hydrolysis for transmembrane transport of a wide range of biomolecules against concentration gradients.

The contribution of methionine to the stability of the Escherichia coli MetNIQ ABC transporter-substrate binding protein complex.

Despite the ubiquitous role of ATP-binding cassette (ABC) importers in nutrient uptake, only the Escherichia coli maltose and vitamin B12 ABC transporters have been structurally characterized in multiple conformations relevant to the alternating access transport mechanism. To complement our previous structure determination of the E. coli MetNI methionine importer in the inward facing conformation (Kadaba et al.

The allosteric regulatory mechanism of the Escherichia coli MetNI methionine ATP binding cassette (ABC) transporter.

The MetNI methionine importer of Escherichia coli, an ATP binding cassette (ABC) transporter, uses the energy of ATP binding and hydrolysis to catalyze the high affinity uptake of D- and L-methionine. Early in vivo studies showed that the uptake of external methionine is repressed by the level of the internal methionine pool, a phenomenon termed transinhibition. Our understanding of the MetNI mechanism has thus far been limited to a series of crystal structures in an inward-facing conformation.

Structural basis for heavy metal detoxification by an Atm1-type ABC exporter.

Although substantial progress has been achieved in the structural analysis of exporters from the superfamily of adenosine triphosphate (ATP)-binding cassette (ABC) transporters, much less is known about how they selectively recognize substrates and how substrate binding is coupled to ATP hydrolysis. We have addressed these questions through crystallographic analysis of the Atm1/ABCB7/HMT1/ABCB6 ortholog from Novosphingobium aromaticivorans DSM 12444, NaAtm1, at 2.4 angstrom resolution.

Dynamics of nucleosome remodelling by individual ACF complexes.

The ATP-dependent chromatin assembly and remodelling factor (ACF) functions to generate regularly spaced nucleosomes, which are required for heritable gene silencing. The mechanism by which ACF mobilizes nucleosomes remains poorly understood. Here we report a single-molecule FRET study that monitors the remodelling of individual nucleosomes by ACF in real time, revealing previously unknown remodelling intermediates and dynamics.

The chromatin remodeller ACF acts as a dimeric motor to space nucleosomes.

Evenly spaced nucleosomes directly correlate with condensed chromatin and gene silencing. The ATP-dependent chromatin assembly factor (ACF) forms such structures in vitro and is required for silencing in vivo. ACF generates and maintains nucleosome spacing by constantly moving a nucleosome towards the longer flanking DNA faster than the shorter flanking DNA.

FRET-based methods to study ATP-dependent changes in chromatin structure.

DNA packaging into chromatin imposes several levels of regulation on the central nuclear processes of DNA replication, recombination, repair and transcription. ATP-dependent chromatin-remodeling enzymes play a critical role in this regulation by altering the accessibility of nucleosomal DNA. Remodeling can result in large-scale changes in chromatin, such as the formation of heterochromatin, or smaller changes in exposure or occlusion of specific DNA regions.

The chromatin-remodeling enzyme ACF is an ATP-dependent DNA length sensor that regulates nucleosome spacing.

Arrays of regularly spaced nucleosomes directly correlate with closed chromatin structures at silenced loci. The ATP-dependent chromatin-assembly factor (ACF) generates such arrays in vitro and is required for transcriptional silencing in vivo. A key unresolved question is how ACF 'measures' equal spacing between nucleosomes.

The rate of bipolar spindle assembly depends on the microtubule-gliding velocity of the mitotic kinesin Eg5.

During early embryonic cycles, the time required for mitotic spindle assembly must match the autonomous cell cycle oscillations because a lack of coordination between these two processes will result in chromosome segregation errors. Members of the widely conserved BimC kinesin family are essential for spindle formation in all eukaryotes, and complete loss of BimC function results in monopolar spindles that have two spindle poles that are not separated. However, the precise roles of BimC motor activity in the spindle assembly process are not known.