Disulfide Cross-linking Reveals a Site of Stable Interaction between C-terminal Regulatory Domains of the Two MalK Subunits in the Maltose Transport Complex

Samanta, Susmita; Ayvaz, Tulin; Reyes, Milagros P.; Shuman, Howard A.; Chen, Jue; Davidson, Amy L.

doi:10.1074/jbc.m301171200

Cited by 35 publications

(42 citation statements)

References 39 publications

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“…The scattered location of the mutations (Table 2) and the EIIA Glc /antibody competition sites made it difficult to propose a binding site for EIIA Glc (66,828). In contrast to Thermococcus litoralis MalK (187), biochemical studies suggest that in the functional MalFGK 2 complex of E. coli, the C termini of the MalK subunits are in close contact (774). This assumption was confirmed by resolving the structures of three different forms of the MalK dimer (117).…”

Section: Inhibits Transcription Induction Mechanismmentioning

confidence: 57%

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

Deutscher

Francke

Postma

2006

Microbiol Mol Biol Rev

1,204

769

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SUMMARY The phosphoenolpyruvate(PEP):carbohydrate phosphotransferase system (PTS) is found only in bacteria, where it catalyzes the transport and phosphorylation of numerous monosaccharides, disaccharides, amino sugars, polyols, and other sugar derivatives. To carry out its catalytic function in sugar transport and phosphorylation, the PTS uses PEP as an energy source and phosphoryl donor. The phosphoryl group of PEP is usually transferred via four distinct proteins (domains) to the transported sugar bound to the respective membrane component(s) (EIIC and EIID) of the PTS. The organization of the PTS as a four-step phosphoryl transfer system, in which all P derivatives exhibit similar energy (phosphorylation occurs at histidyl or cysteyl residues), is surprising, as a single protein (or domain) coupling energy transfer and sugar phosphorylation would be sufficient for PTS function. A possible explanation for the complexity of the PTS was provided by the discovery that the PTS also carries out numerous regulatory functions. Depending on their phosphorylation state, the four proteins (domains) forming the PTS phosphorylation cascade (EI, HPr, EIIA, and EIIB) can phosphorylate or interact with numerous non-PTS proteins and thereby regulate their activity. In addition, in certain bacteria, one of the PTS components (HPr) is phosphorylated by ATP at a seryl residue, which increases the complexity of PTS-mediated regulation. In this review, we try to summarize the known protein phosphorylation-related regulatory functions of the PTS. As we shall see, the PTS regulation network not only controls carbohydrate uptake and metabolism but also interferes with the utilization of nitrogen and phosphorus and the virulence of certain pathogens.

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Section: Inhibits Transcription Induction Mechanismmentioning

confidence: 57%

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

Deutscher

Francke

Postma

2006

Microbiol Mol Biol Rev

1,204

769

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“…This intrinsic mobility of the nucleotide-free form of NBDs is further reflected in the crystal structures of E. coli MalK . MalK contains an additional regulatory domain that enhances 'dimer' formation (Samanta et al 2003), thus producing 'dimers' of the apo form of MalK. Here, the authors present two different dimer conformations, accounting for different intermediate forms of the nucleotide-free protein.…”

Section: The Intrinsic Flexibility Of the Apo Nbd Formsmentioning

confidence: 98%

The motor domains of ABC-transporters

Oswald

Holland

Schmitt

2006

Naunyn Schmied Arch Pharmacol

133

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The transport of substrates across a cellular membrane is a vitally important biological function essential for cell survival. ATP-binding cassette (ABC) transporters constitute one of the largest subfamilies of membrane proteins, accomplishing this task. Mutations in genes encoding for ABC transporters cause different diseases, for example, Adrenoleukodystrophy, Stargardt disease or Cystic Fibrosis. Furthermore, some ABC transporters are responsible for multidrug resistance, presenting a major obstacle in modern cancer chemotherapy. In order to translocate the enormous variety of substrates, ranging from ions, nutrients, small peptides to large toxins, different ABC-transporters utilize the energy gained from ATP binding and hydrolysis. The ATP binding cassette, also called the motor domain of ABC transporters, is highly conserved among all ABC transporters. The ability to purify this domain rather easily presents a perfect possibility to investigate the mechanism of ATP hydrolysis, thus providing us with a detailed picture of this process. Recently, many crystal structures of the ATP-binding domain and the full-length structures of two ABC transporters have been solved. Combining these structural data, we have now the opportunity to analyze the hydrolysis event on a molecular level. This review provides an overview of the structural investigations of the ATPbinding domains, highlighting molecular changes upon ATP binding and hydrolysis.

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“…Precedent for interacting proteins affecting transport exists. In particular, MalK has been shown to interact with EIIA Glc (45) and the maltose regulator MalT (13,46). Our work is now focusing on identifying proteins that may interact with RhaK to affect rhamnose transport.…”

Section: Discussionmentioning

confidence: 99%

Carbohydrate Kinase (RhaK)-Dependent ABC Transport of Rhamnose in Rhizobium leguminosarum Demonstrates Genetic Separation of Kinase and Transport Activities

Rivers

Oresnik

2013

J Bacteriol

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In Rhizobium leguminosarum the ABC transporter responsible for rhamnose transport is dependent on RhaK, a sugar kinase that is necessary for the catabolism of rhamnose. This has led to a working hypothesis that RhaK has two biochemical functions: phosphorylation of its substrate and affecting the activity of the rhamnose ABC transporter. To address this hypothesis, a linkerscanning random mutagenesis of rhaK was carried out. Thirty-nine linker-scanning mutations were generated and mapped. Alleles were then systematically tested for their ability to physiologically complement kinase and transport activity in a strain carrying an rhaK mutation. The rhaK alleles generated could be divided into three classes: mutations that did not affect either kinase or transport activity, mutations that eliminated both transport and kinase activity, and mutations that affected transport activity but not kinase activity. Two genes of the last class (rhaK72 and rhaK73) were found to have similar biochemical phenotypes but manifested different physiological phenotypes. Whereas rhaK72 conferred a slow-growth phenotype when used to complement rhaK mutants, the rhaK73 allele did not complement the inability to use rhamnose as a sole carbon source. To provide insight to how these insertional variants might be affecting rhamnose transport and catabolism, structural models of RhaK were generated based on the crystal structure of related sugar kinases. Structural modeling suggests that both rhaK72 and rhaK73 affect surface-exposed residues in two distinct regions that are found on one face of the protein, suggesting that this protein's face may play a role in protein-protein interaction that affects rhamnose transport. C ells require a specific and regulated way to transport substrates across membranes. One of the largest families of transporters is the ATP binding cassette (ABC) transporters. ABC transporters utilize free energy from ATP hydrolysis to transport substrates across a membrane. They are widely distributed in all domains of life and are involved in transport that affects diverse biological functions (1-3).Members of the ABC superfamily are defined by the ATPase protein that contains the Walker A and Walker B motifs, along with an LSGGQ conserved consensus sequence (4, 5). Functional ABC importers generally consist of two proteins that have transmembrane domains consisting of six membrane-spanning regions (permeases), two ABC proteins that are cytoplasmically localized and contain the ATP binding domains, and a periplasmically localized substrate binding protein for the function of the transport system (6). Relatively few sequence similarities are found between binding proteins for different substrates as the periplasmic substrate binding protein plays a central role in substrate specificity (7).Gram-negative bacterial ABC transport systems responsible for the import of carbohydrates can be broken into two main classes: carbohydrate uptake transporters 1 and 2 (CUT1 and -2, respectively) (8). Most work on bacterial carbohydrate...

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Disulfide Cross-linking Reveals a Site of Stable Interaction between C-terminal Regulatory Domains of the Two MalK Subunits in the Maltose Transport Complex

Cited by 35 publications

References 39 publications

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

The motor domains of ABC-transporters

Carbohydrate Kinase (RhaK)-Dependent ABC Transport of Rhamnose in Rhizobium leguminosarum Demonstrates Genetic Separation of Kinase and Transport Activities

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