scholarly journals Functional Determinants of Metal Ion Transport and Selectivity in Paralogous Cation Diffusion Facilitator Transporters CzcD and MntE in Streptococcus pneumoniae

2016 ◽  
Vol 198 (7) ◽  
pp. 1066-1076 ◽  
Author(s):  
Julia E. Martin ◽  
David P. Giedroc
Keyword(s):  
Streptococcus Pneumoniae ◽  
Metal Ion ◽  
Transmembrane Domain ◽  
Divalent Metal ◽  
Cation Diffusion ◽  
Metal Specificity ◽  
Cation Diffusion Facilitator ◽  
Content Type ◽  
Metal Selectivity ◽  
A Site

ABSTRACTCation diffusion facilitators (CDFs) are a large family of divalent metal transporters that collectively possess broad metal specificity and contribute to intracellular metal homeostasis and virulence in bacterial pathogens.Streptococcus pneumoniaeexpresses two homologous CDF efflux transporters, MntE and CzcD. Cells lackingmntEorczcDare sensitive to manganese (Mn) or zinc (Zn) toxicity, respectively, and specifically accumulate Mn or Zn, respectively, thus suggesting that MntE selectively transports Mn, while CzcD transports Zn. Here, we probe the origin of this metal specificity using a phenotypic growth analysis of pneumococcal variants. Structural homology toEscherichia coliYiiP predicts that both MntE and CzcD are dimeric and each protomer harbors four pairs of conserved metal-binding sites, termed the A site, the B site, and the C1/C2 binuclear site. We find that single amino acid mutations within both the transmembrane domain A site and the B site in both CDFs result in a cellular metal sensitivity similar to that of the corresponding null mutants. However, multiple mutations in the predicted cytoplasmic C1/C2 cluster of MntE have no impact on cellular Mn resistance, in contrast to the analogous substitutions in CzcD, which do have on impact on cellular Zn resistance. Deletion of the MntE-specific C-terminal tail, present only in Mn-specific bacterial CDFs, resulted in only a modest growth phenotype. Further analysis of MntE-CzcD functional chimeric transporters showed that Asn and Asp in theND-DD A-site motif of MntE and the most N-terminal His in theHD-HD site A of CzcD (the specified amino acids are underlined) play key roles in transporter metal selectivity.IMPORTANCECation diffusion facilitator (CDF) proteins are divalent metal ion transporters that are conserved in organisms ranging from bacteria to humans and that play important roles in cellular physiology, from metal homeostasis and resistance to type I diabetes in vertebrates. The respiratory pathogenStreptococcus pneumoniaeexpresses two metal CDF transporters, CzcD and MntE. How CDFs achieve metal selectivity is unclear. We show here that CzcD and MntE are true paralogs, as CzcD transports zinc, while MntE selectively transports manganese. Through the use of an extensive collection of pneumococcal variants, we show that a primary determinant for metal selectivity is the A site within the transmembrane domain. This extends our understanding of how CDFs discriminate among transition metals.

scholarly journals Multi-genomic analysis of the Cation Diffusion Facilitator transporters from algae

2019 ◽  
Author(s):  
Aniefon Ibuot ◽  
Andrew P. Dean ◽  
Jon K. Pittman
Keyword(s):  
Metal Ion ◽  
Transmembrane Domain ◽  
Higher Plants ◽  
Phylogenetic Analyses ◽  
Algal Species ◽  
Genomic Analysis ◽  
Transport Processes ◽  
Transport Function ◽  
Cation Diffusion ◽  
Cation Diffusion Facilitator

AbstractMetal transport processes are relatively poorly understood in algae in comparison to higher plants and other eukaryotes. A screen of genomes from 33 taxonomically diverse algal species was conducted to identify members of the Cation Diffusion Facilitator (CDF) family of metal ion transporter. All algal genomes contained at least one CDF gene with four species having >10 CDF genes (median of 5 genes per genome), further confirming that this is a ubiquitous gene family. Phylogenetic analysis suggested a CDF gene organisation of five groups, which includes Zn-CDF, Fe/Zn-CDF and Mn-CDF groups, consistent with previous phylogenetic analyses, and two functionally undefined groups. One of these undefined groups was algal specific although excluded chlorophyte and rhodophyte sequences. The majority of sequences (22 out of 26 sequences) from this group had a putative ion binding site motif within transmembrane domain 2 and 5 that was distinct from other CDF proteins, such that alanine or serine replaced the conserved histidine residue. The phylogenetic grouping was supported by sequence cluster analysis. Yeast heterologous expression of CDF proteins from Chlamydomonas reinhardtii indicated Zn2+ and Co2+ transport function by CrMTP1, and Mn2+ transport function by CrMTP2, CrMTP3 and CrMTP4, which validated the phylogenetic prediction. However, the Mn-CDF protein CrMTP3 was also able to provide zinc and cobalt tolerance to the Zn- and Co-sensitive zrc1cot1 yeast strain. There is wide diversity of CDF transporters within the algae lineage, and some of these genes may be attractive targets for future applications of metal content engineering in plants or microorganisms.

scholarly journals The cation diffusion facilitator protein MamM's cytoplasmic domain exhibits metal-type dependent binding modes and discriminates against Mn2+

2020 ◽  
Vol 295 (49) ◽  
pp. 16614-16629
Author(s):  
Shiran Barber-Zucker ◽  
Jenny Hall ◽  
Afonso Froes ◽  
Sofiya Kolusheva ◽  
Fraser MacMillan ◽  
...  
Keyword(s):  
Transmembrane Domain ◽  
Magnetotactic Bacteria ◽  
Crystal Form ◽  
Binding Modes ◽  
Cation Diffusion ◽  
Cation Diffusion Facilitator ◽  
Metal Type ◽  
Metal Selectivity ◽  
Functional Discrimination ◽  
Metal Recognition

Cation diffusion facilitator (CDF) proteins are a conserved family of divalent transition metal cation transporters. CDF proteins are usually composed of two domains: the transmembrane domain, in which the metal cations are transported through, and a regulatory cytoplasmic C-terminal domain (CTD). Each CDF protein transports either one specific metal or multiple metals from the cytoplasm, and it is not known whether the CTD takes an active regulatory role in metal recognition and discrimination during cation transport. Here, the model CDF protein MamM, an iron transporter from magnetotactic bacteria, was used to probe the role of the CTD in metal recognition and selectivity. Using a combination of biophysical and structural approaches, the binding of different metals to MamM CTD was characterized. Results reveal that different metals bind distinctively to MamM CTD in terms of their binding sites, thermodynamics, and binding-dependent conformations, both in crystal form and in solution, which suggests a varying level of functional discrimination between CDF domains. Furthermore, these results provide the first direct evidence that CDF CTDs play a role in metal selectivity. We demonstrate that MamM's CTD can discriminate against Mn2+, supporting its postulated role in preventing magnetite formation poisoning in magnetotactic bacteria via Mn2+ incorporation.

scholarly journals The cation diffusion facilitator protein MamM’s cytoplasmic domain exhibits metal-type dependent binding modes and discriminates against Mn2+

2020 ◽  
Author(s):  
Shiran Barber-Zucker ◽  
Jenny Hall ◽  
Afonso Froes ◽  
Sofiya Kolusheva ◽  
Fraser MacMillan ◽  
...  
Keyword(s):  
Direct Evidence ◽  
Transmembrane Domain ◽  
Crystal Form ◽  
Binding Modes ◽  
Cation Diffusion ◽  
Cation Diffusion Facilitator ◽  
Metal Type ◽  
Metal Selectivity ◽  
Specific Metal

SummaryCation diffusion facilitator (CDF) proteins are a conserved family of divalent transition metal cation transporters. CDF proteins are usually composed of two domains: the transmembrane domain (TMD), in which the metal cations are transported through, and a regulatory cytoplasmic C-terminal domain (CTD). Each CDF protein transports either one specific metal, or multiple metals, from the cytoplasm. Here, the model CDF protein MamM, from magnetotactic bacteria, was used to probe the role of the CTD in metal selectivity. Using a combination of biophysical and structural approaches, the binding of different metals to MamM CTD was characterized. Results reveal that different metals bind distinctively to MamM CTD in terms of; their binding sites, thermodynamics and binding-dependent conformation, both in crystal form and in solution. Furthermore, the results indicate that the CTD discriminates against Mn2+ and provides the first direct evidence that CDF CTD’s play a role in metal selectivity.

scholarly journals The metal binding site composition of the cation diffusion facilitator protein MamM cytoplasmic domain impacts its metal responsivity

2020 ◽  
Author(s):  
Shiran Barber-Zucker ◽  
Anat Shahar ◽  
Sofiya Kolusheva ◽  
Raz Zarivach
Keyword(s):  
Metal Binding ◽  
Binding Sites ◽  
Transmembrane Domain ◽  
Metal Cations ◽  
Data Bank ◽  
Cation Diffusion ◽  
Cation Diffusion Facilitator ◽  
Metal Selectivity ◽  
The Impact

AbstractThe cation diffusion facilitator (CDF) is a conserved family of divalent d-block metal cation transporters that extrude these cations selectively from the cytoplasm. CDF proteins are composed of two domains: the transmembrane domain, through which the cations are transported, and a regulatory cytoplasmic C-terminal domain (CTD). Metal binding to the CTD leads to its tighter conformation, and this sequentially promotes conformational change of the transmembrane domain which allows the actual transport of specific metal cations. It was recently shown that the magnetotactic bacterial CDF protein MamM CTD has a role in metal selectivity, as binding of different metal cations exhibits distinctive affinities and conformations. It is yet unclear whether the composition of the CTD binding sites can impact metal selectivity. Here we performed a mutational study of MamM CTD, where we exchanged the metal binding residues with different metal-binding amino acids. Using X-ray crystallography and Trp-fluorescence spectrometry, we studied the impact of the mutations on the CTD conformation in the presence of different metals. Our results reveal that the incorporation of such mutations alters the domain response to metals in vitro, as mutant forms of the CTD bind metals differently in terms of the composition of the binding sites and the CTD conformation.CoordinatesMamM CTD structures have been deposited in the Protein Data Bank under the following accession codes: 6H5V, 6H5M, 6H5U, 6H8G, 6HAO, 6H88, 6H87, 6H8A, 6H89, 6H8D, 6H5K, 6H9Q, 6H84, 6H83, 6HA2, 6H8I, 6H9T, 6H81, 6HAN, 6H85, 6H9P, 6HHS.

A Preliminary Bioinformatics Analysis of Cation Diffusion Facilitator (CDF) Proteins in Different Plants

2011 ◽  
Vol 282-283 ◽  
pp. 453-456
Author(s):  
Hua Zhong ◽  
Bao Ping Sun ◽  
Fang Ying Zhao
Keyword(s):  
Secretory Pathway ◽  
Transmembrane Domain ◽  
Transmembrane Protein ◽  
Populus Trichocarpa ◽  
Alpha Helix ◽  
Random Coil ◽  
Post Translational Modification ◽  
Cation Diffusion ◽  
Cation Diffusion Facilitator ◽  
Arabidopsis Lyrata

The amino acid sequence of Cation Diffusion Facilitator from Populus trichocarpa, Thlaspi goesingense, Arabidopsis lyrata subsp. Lyrata, Brassica juncea and Medicago sativa, registered in GenBank, were analyzed and researched by the bioinformatic tools in the several aspects, including hydrophobicity / hydrophilicity properties, post-translational modification, secondary structures prediction and transmembrane domain. The results showed that Cation Diffusion Facilitator is a hydrophobic and transmembrane protein, which exists in endoplasmic reticulum and other secretory pathway. The main motifs of predicted secondary structure of Cation Diffusion Facilitator are alpha helix and random coil.

scholarly journals Manganese Detoxification by MntE Is Critical for Resistance to Oxidative Stress and Virulence of Staphylococcus aureus

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Caroline M. Grunenwald ◽  
Jacob E. Choby ◽  
Lillian J. Juttukonda ◽  
William N. Beavers ◽  
Andy Weiss ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Staphylococcus Aureus ◽  
Pathogenic Bacteria ◽  
Efflux Pump ◽  
Uptake System ◽  
Cation Diffusion ◽  
Intracellular Iron ◽  
Cation Diffusion Facilitator ◽  
Mn Toxicity ◽  
Content Type

ABSTRACT Manganese (Mn) is an essential micronutrient critical for the pathogenesis of Staphylococcus aureus, a significant cause of human morbidity and mortality. Paradoxically, excess Mn is toxic; therefore, maintenance of intracellular Mn homeostasis is required for survival. Here we describe a Mn exporter in S. aureus, MntE, which is a member of the cation diffusion facilitator (CDF) protein family and conserved among Gram-positive pathogens. Upregulation of mntE transcription in response to excess Mn is dependent on the presence of MntR, a transcriptional repressor of the mntABC Mn uptake system. Inactivation of mntE or mntR leads to reduced growth in media supplemented with Mn, demonstrating MntE is required for detoxification of excess Mn. Inactivation of mntE results in elevated levels of intracellular Mn, but reduced intracellular iron (Fe) levels, supporting the hypothesis that MntE functions as a Mn efflux pump and Mn efflux influences Fe homeostasis. Strains inactivated for mntE are more sensitive to the oxidants NaOCl and paraquat, indicating Mn homeostasis is critical for resisting oxidative stress. Furthermore, mntE and mntR are required for full virulence of S. aureus during infection, suggesting S. aureus experiences Mn toxicity in vivo. Combined, these data support a model in which MntR controls Mn homeostasis by balancing transcriptional repression of mntABC and induction of mntE, both of which are critical for S. aureus pathogenesis. Thus, Mn efflux contributes to bacterial survival and virulence during infection, establishing MntE as a potential antimicrobial target and expanding our understanding of Mn homeostasis. IMPORTANCE Manganese (Mn) is generally viewed as a critical nutrient that is beneficial to pathogenic bacteria due to its function as an enzymatic cofactor and its capability of acting as an antioxidant; yet paradoxically, high concentrations of this transition metal can be toxic. In this work, we demonstrate Staphylococcus aureus utilizes the cation diffusion facilitator (CDF) family protein MntE to alleviate Mn toxicity through efflux of excess Mn. Inactivation of mntE leads to a significant reduction in S. aureus resistance to oxidative stress and S. aureus-mediated mortality within a mouse model of systemic infection. These results highlight the importance of MntE-mediated Mn detoxification in intracellular Mn homeostasis, resistance to oxidative stress, and S. aureus virulence. Therefore, this establishes MntE as a potential target for development of anti-S. aureus therapeutics.

scholarly journals Putative metal binding site in the transmembrane domain of the manganese transporter SLC30A10 is different from that of related zinc transporters

Metallomics ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 1053-1064 ◽  
Author(s):  
Charles E. Zogzas ◽  
Somshuvra Mukhopadhyay
Keyword(s):  
Binding Site ◽  
Metal Binding ◽  
Transmembrane Domain ◽  
Zinc Transporters ◽  
Cation Diffusion ◽  
Metal Binding Site ◽  
Cation Diffusion Facilitator

Mechanism by which the cation diffusion facilitator SLC30A10 transports manganese is fundamentally different from that of previously-studied proteins in this superfamily.

scholarly journals An Electrostatic Interaction between BlpC and BlpH Dictates Pheromone Specificity in the Control of Bacteriocin Production and Immunity in Streptococcus pneumoniae

2015 ◽  
Vol 197 (7) ◽  
pp. 1236-1248 ◽  
Author(s):  
Marisa D. Pinchas ◽  
Nathan C. LaCross ◽  
Suzanne Dawid
Keyword(s):  
Streptococcus Pneumoniae ◽  
Quorum Sensing ◽  
Electrostatic Interaction ◽  
Transmembrane Domain ◽  
Allelic Variation ◽  
Single Species ◽  
Bacteriocin Production ◽  
Content Type ◽  
Naturally Occurring ◽  
Pheromone Specificity

ABSTRACTTheblplocus ofStreptococcus pneumoniaesecretes and regulates bacteriocins, which mediate both intra- and interspecific competition in the human nasopharynx. There are four major alleles of the geneblpH, which encodes the receptor responsible for activating theblplocus when bound to one of four distinct peptide pheromones (BlpC). The allelic variation ofblpHis presumably explained by a need to restrict cross talk between competing strains. The BlpH protein sequences have polymorphisms distributed throughout the sequence, making identification of the peptide binding site difficult to predict. To identify the pheromone binding sites that dictate pheromone specificity, we have characterized the four major variants and two naturally occurring chimeric versions ofblpHin which recombination events appear to have joined two distinctblpHalleles together. Using these allelic variants, a series of laboratory-generated chimericblpHalleles, and site-directed mutants of both the receptor and peptide, we have demonstrated that BlpC binding to some BlpH types involves an electrostatic interaction between the oppositely charged residues of BlpC and the first transmembrane domain of BlpH. An additional recognition site was identified in the second extracellular loop. We identified naturally occurring BlpH types that have the capacity to respond to more than one BlpC type; however, this change in specificity results in a commensurate drop in overall sensitivity. These natural recombination events were presumably selected for to balance the need to sense bacteriocin-secreting neighbors with the need to turn on bacteriocin production at a low density.IMPORTANCEBacteria use quorum sensing to optimize gene expression to accommodate for local bacterial density and diffusion rates. To prevent interception of quorum-sensing signals by neighboring strains, the genomes of single species often encode strain-specific signal/receptor pairs. Theblplocus inStreptococcus pneumoniaethat drives bacteriocin secretion is controlled by quorum sensing that involves the interaction of the signal/receptor pair BlpC/BlpH. We show that the pneumococcal population can be divided into several distinct BlpC/BlpH pairs; however, there are examples of naturally occurring chimeric receptors that can bind to more than one BlpC type. The trade-off for this broadened specificity is a loss of overall receptor sensitivity. This suggests that under certain conditions, the advantage of signal interception can trump the requirements for self-induction.

scholarly journals Transition metal binding selectivity in proteins and its correlation with the phylogenomic classification of the cation diffusion facilitator protein family

2017 ◽  
Author(s):  
Shiran Barber-Zucker ◽  
Boaz Shaanan ◽  
Raz Zarivach
Keyword(s):  
Binding Site ◽  
De Novo ◽  
Specific Binding ◽  
Protein Family ◽  
Cation Diffusion ◽  
Binding Selectivity ◽  
Cation Diffusion Facilitator ◽  
Conserved Sequence ◽  
Metal Selectivity

AbstractDivalent d-block metal cations (DDMCs), such as Fe, Zn and Mn, participate in many biological processes. Understanding how specific DDMCs are transported to and within the cell and what controls their binding selectivity to different proteins is crucial for defining the mechanisms of metalloproteins. To better understand such processes, we scanned the RCSB Protein Data Bank, performed a de novo structural-based comprehensive analysis of seven DDMCs and found their amino acid binding and coordination geometry propensities. We then utilized these results to characterize the correlation between metal selectivity, specific binding site composition and phylogenetic classification of the cation diffusion facilitator (CDF) protein family, a family of DDMC transporters found throughout evolution and sharing a conserved structure, yet with different members displaying distinct metal selectivity. Our analysis shows that DDMCs differ, at times significantly, in terms of their binding propensities, and that in each CDF clade, the metal selectivity-related binding site has a unique and conserved sequence signature. However, only limited correlation exists between the composition of the DDMC binding site in each clade and the metal selectivity shown by its proteins.

Sign in / Sign up

Export Citation Format

Share Document