Journal of Molecular Biology

ScienceDirect RSS
  • A Mutation in the Catalytic Loop of Hsp90 Specifically Impairs ATPase Stimulation by Aha1p, But Not Hch1p
    [Jun 2014]

    Publication date: 12 June 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 12

    Author(s): Natalie K. Horvat , Heather Armstrong , Brian L. Lee , Rebecca Mercier , Annemarie Wolmarans , Jacob Knowles , Leo Spyracopoulos , Paul LaPointe

    Heat shock protein 90 (Hsp90) is a molecular chaperone that plays a central role in maintaining cellular homeostasis by facilitating activation of a large number of client proteins. ATP-dependent client activation by Hsp90 is tightly regulated by a host of co-chaperone proteins that control progression through the activation cycle. ATPase stimulation of Hsp90 by Aha1p requires a conserved RKxK motif that interacts with the catalytic loop of Hsp90. In this study, we explore the role of this RKxK motif in the biological and biochemical properties of Hch1p. We found that this motif is required for Hch1p-mediated ATPase stimulation in vitro, but mutations that block stimulation do not impair the action of Hch1p in vivo. This suggests that the biological function of Hch1p is not directly linked to ATPase stimulation. Moreover, a mutation in the catalytic loop of Hsp90 specifically impairs ATPase stimulation by Aha1p but not by Hch1p. Our work here suggests that both Hch1p and Aha1p regulate Hsp90 function through interaction with the catalytic loop but do so in different ways.
    Graphical abstract




    Categories: Journal Articles
  • Elucidating the Mechanism of Substrate Recognition by the Bacterial Hsp90 Molecular Chaperone
    [Jun 2014]

    Publication date: 12 June 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 12

    Author(s): Timothy O. Street , Xiaohui Zeng , Riccardo Pellarin , Massimiliano Bonomi , Andrej Sali , Mark J.S. Kelly , Feixia Chu , David A. Agard

    Hsp90 is a conformationally dynamic molecular chaperone known to promote the folding and activation of a broad array of protein substrates (“clients”). Hsp90 is believed to preferentially interact with partially folded substrates, and it has been hypothesized that the chaperone can significantly alter substrate structure as a mechanism to alter the substrate functional state. However, critically testing the mechanism of substrate recognition and remodeling by Hsp90 has been challenging. Using a partially folded protein as a model system, we find that the bacterial Hsp90 adapts its conformation to the substrate, forming a binding site that spans the middle and C-terminal domains of the chaperone. Cross-linking and NMR measurements indicate that Hsp90 binds to a large partially folded region of the substrate and significantly alters both its local and long-range structure. These findings implicate Hsp90's conformational dynamics in its ability to bind and remodel partially folded proteins. Moreover, native-state hydrogen exchange indicates that Hsp90 can also interact with partially folded states only transiently populated from within a thermodynamically stable, native-state ensemble. These results suggest a general mechanism by which Hsp90 can recognize and remodel native proteins by binding and remodeling partially folded states that are transiently sampled from within the native ensemble.
    Graphical abstract




    Categories: Journal Articles
  • A Genome-Wide Sequence–Structure Analysis Suggests Aggregation Gatekeepers Constitute an Evolutionary Constrained Functional Class
    [Jun 2014]

    Publication date: 12 June 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 12

    Author(s): Greet De Baets , Joost Van Durme , Frederic Rousseau , Joost Schymkowitz

    Protein aggregation is geared by aggregation-prone regions that self-associate by β-strand interactions. Charged residues and prolines are enriched at the flanks of aggregation-prone regions resulting in decreased aggregation. It is still unclear what drives the overrepresentation of these “aggregation gatekeepers”, that is, whether their presence results from structural constraints determining protein stability or whether they constitute a bona fide functional class selectively maintained to control protein aggregation. As functional residues are typically conserved regardless of their cost to protein stability, we compared sequence conservation and thermodynamic cost of these residues in 2659 protein families in Escherichia coli. Across protein families, we find gatekeepers to be under strong selective conservation while at the same time representing a significant thermodynamic cost to protein structure. This finding supports the notion that aggregation gatekeepers are not structurally determined but evolutionary selected to control protein aggregation.
    Graphical abstract




    Categories: Journal Articles
  • The positive inside rule is stronger when followed by a transmembrane helix
    [Jun 2014]

    Publication date: Available online 10 June 2014
    Source:Journal of Molecular Biology

    Author(s): Minttu T. Virkki , Christoph Peters , Daniel Nilsson , Therese Sörensen , Susana Cristobal , Björn Wallner , Arne Elofsson

    The translocon recognizes transmembrane helices with sufficient level of hydrophobicity and inserts them into the membrane. However, sometimes less hydrophobic helices are also recognized. Positive inside rule, orientational preferences of and specific interactions with neighboring helices have been shown to aid in the recognition of these helices, at least in artificial systems. To better understand how the translocon inserts marginally hydrophobic helices, we studied three naturally occurring marginally hydrophobic helices, which were previously shown to require the subsequent helix for efficient translocon recognition. We find no evidence for specific interactions when we scan all residues in the subsequent helices. Instead, we identify arginines located at the N-terminal part of the subsequent helices that are crucial for the recognition of the marginally hydrophobic transmembrane helices, indicating that the positive inside rule is important. However, in two of the constructs these arginines do not aid in the recognition without the rest of the subsequent helix, i.e. the positive inside rule alone is not sufficient. Instead, the improved recognition of marginally hydrophobic helices can here be explained as follows; the positive inside rule provides an orientational preference of the subsequent helix, which in turn allows the marginally hydrophobic helix to be inserted, i.e. the effect of the positive inside rule is stronger if positively charged residues are followed by a transmembrane helix. Such a mechanism can obviously not aid C-terminal helices and consequently we find that the terminal helices in multi-spanning membrane proteins are more hydrophobic than internal helices.
    Graphical abstract




    Categories: Journal Articles
  • Sus1p facilitates pre-initiation complex formation at the SAGA-regulated genes independently of histone H2B de-ubiquitylation
    [Jun 2014]

    Publication date: Available online 6 June 2014
    Source:Journal of Molecular Biology

    Author(s): Geetha Durairaj , Rwik Sen , Bhawana Uprety , Abhijit Shukla , Sukesh R. Bhaumik

    Sus1p is a common component of transcriptional co-activator, SAGA (Spt-Ada-Gcn5-Acetyltransferase), and mRNA export complex, TREX-2 (Transcription-export 2), and is involved in promoting transcription as well as mRNA export. However, it is not clearly understood how Sus1p promotes transcription. Here, we show that Sus1p is predominantly recruited to the upstream activating sequence of a SAGA-dependent gene, GAL1, under transcriptionally active conditions as a component of SAGA to promote the formation of pre-initiation complex (PIC) at the core promoter, and consequently, transcriptional initiation. Likewise, Sus1p promotes the PIC formation at other SAGA-dependent genes, and hence transcriptional initiation. Such function of Sus1p in promoting PIC formation and transcriptional initiation is not mediated via its role in regulation of SAGA’s histone H2B de-ubiquitylation activity. However, Sus1p’s function in regulation of histone H2B ubiquitylation is associated with transcriptional elongation, DNA repair and replication. Collectively, our results support that Sus1p promotes PIC formation (and hence transcriptional initiation) at the SAGA-regulated genes independently of histone H2B de-ubiquitylation, and further controls transcriptional elongation, DNA repair and replication via orchestration of histone H2B ubiquitylation, thus providing distinct functional insights of Sus1p in regulation of DNA transacting processes.
    Graphical abstract




    Categories: Journal Articles
  • The Yin and Yang of Bacterial Resilience in the Human Gut Microbiota
    [Jun 2014]

    Publication date: Available online 6 June 2014
    Source:Journal of Molecular Biology

    Author(s): Molly K. Gibson , Mitchell W. Pesesky , Gautam Dantas

    The human gut is home to trillions of microbes that form a symbiotic relationship with the human host. During health, the intestinal microbiota provides many benefits to the host and is generally resistant to colonization by new species; however, disruption of this complex community can lead to pathogen invasion, inflammation, and disease. Restoration and maintenance of a healthy gut microbiota composition requires effective therapies to reduce and prevent colonization of harmful bacteria (pathogens) while simultaneously promoting growth of beneficial bacteria (probiotics). Here we review the mechanisms by which the host modulates the gut community composition during health and disease, and discuss prospects for antibiotic and probiotic therapy for restoration of a healthy intestinal community following disruption.
    Graphical abstract




    Categories: Journal Articles
  • Chaperonins Resculpt Folding Free Energy Landscapes To Avoid Kinetic Traps and Accelerate Protein Folding
    [Jun 2014]

    Publication date: Available online 5 June 2014
    Source:Journal of Molecular Biology

    Author(s): Xin Zhang , Jeffery W. Kelly







    Categories: Journal Articles
  • Crystal Structures of Ricin Toxin’s Enzymatic Subunit (RTA) in Complex with Neutralizing and Non-neutralizing Single Chain Antibodies
    [Jun 2014]

    Publication date: Available online 4 June 2014
    Source:Journal of Molecular Biology

    Author(s): Michael J. Rudolph , David J. Vance , Jonah Cheung , Matthew C. Franklin , Fiana Burshteyn , Michael S. Cassidy , Ebony N. Gary , Cristina Herrera , Charles B. Shoemaker , Nicholas J. Mantis

    Ricin is a Select Agent Toxin and a member of the RNA N-glycosidase family of medically important plant and bacterial ribosome-inactivating proteins (RIPs). In this study, we determined x-ray crystal structures of the enzymatic subunit of ricin (RTA) in complex with the antigen binding domains (VHH) of five unique single-chain monoclonal antibodies that differ in their respective toxin-neutralizing activities. None of the VHHs made direct contact with residues involved in RTA’s RNA N-glycosidase activity or induced notable allosteric changes in the toxin’s subunit. Rather, the five VHHs had overlapping structural epitopes on the surface of the toxin and differed in the degree to which they made contact with prominent structural elements in two folding domains of the RTA. In general, RTA interactions were influenced most by the VHH CDR3 elements, with the most potent neutralizing antibody having the shortest and most conformationally constrained CDR3. These structures provide unique insights into the mechanisms underlying toxin neutralization and provide critically important information required for the rational design of ricin toxin subunit vaccines.
    Graphical abstract




    Categories: Journal Articles
  • Chromatin Structure and Replication Origins: Determinants Of Chromosome Replication And Nuclear Organization
    [Jun 2014]

    Publication date: Available online 3 June 2014
    Source:Journal of Molecular Biology

    Author(s): Owen K. Smith , Mirit I. Aladjem

    The DNA replication program is, in part, determined by the epigenetic landscape that governs local chromosome architecture and directs chromosome duplication. Replication must coordinate with other biochemical processes occurring concomitantly on chromatin, such as transcription and remodeling, to insure accurate duplication of both genetic and epigenetic features and to preserve genomic stability. The importance of genome architecture and chromatin looping in coordinating cellular processes on chromatin is illustrated by two recent sets of discoveries. First, chromatin-associated proteins that are not part of the core replication machinery were shown to affect the timing of DNA replication. These chromatin-associated proteins could be working in concert, or perhaps in competition, with the transcriptional machinery and with chromatin modifiers to determine the spatial and temporal organization of replication initiation events. Second, epigenetic interactions are mediated by DNA sequences that determine chromosomal replication. In this review we summarize recent findings and current models linking spatial and temporal regulation of the replication program with epigenetic signaling. We discuss these issues in the context of the genome’s three-dimensional structure with an emphasis on events occurring during the initiation of DNA replication.
    Graphical abstract




    Categories: Journal Articles
  • Reshaping chromatin after DNA damage: the choreography of histone proteins
    [Jun 2014]

    Publication date: Available online 1 June 2014
    Source:Journal of Molecular Biology

    Author(s): Sophie E. Polo

    DNA damage signaling and repair machineries operate in a nuclear environment, where DNA is wrapped around histone proteins and packaged into chromatin. Understanding how chromatin structure is restored together with the DNA sequence during DNA damage repair has been a topic of intense research. Indeed, chromatin integrity is central to cell functions and identity. Yet, chromatin shows remarkable plasticity in response to DNA damage. This review presents our current knowledge of chromatin dynamics in the mammalian cell nucleus in response to DNA-double strand breaks and UV lesions. I provide an overview of the key players involved in regulating histone dynamics in damaged chromatin regions, focusing on histone chaperones and their concerted action with histone modifiers, chromatin remodelers and repair factors. I also discuss how these dynamics contribute to reshaping chromatin and, by altering the chromatin landscape, may affect the maintenance of epigenetic information.
    Graphical abstract




    Categories: Journal Articles
  • Pyruvate Formate-Lyase Interacts Directly with the Formate Channel FocA to Regulate Formate Translocation
    [Jun 2014]

    Publication date: Available online 1 June 2014
    Source:Journal of Molecular Biology

    Author(s): Claudia Doberenz , Michael Zorn , Dörte Falke , David Nannemann , Doreen Hunger , Lydia Beyer , Christian H. Ihling , Jens Meiler , Andrea Sinz , R. Gary Sawers

    The formate-nitrite transporters (FNT) form a superfamily of pentameric membrane channels that translocate monovalent anions across biological membranes. FocA translocates formate bidirectionally but the mechanism underlying how translocation of formate is controlled and what governs substrate specificity remain unclear. Here we demonstrate that the normally soluble dimeric enzyme pyruvate formate-lyase (PflB), which is responsible for intracellular formate generation in enterobacteria and other microbes, interacts specifically with FocA. Association of PflB with the cytoplasmic membrane was shown to be FocA-dependent and purified, Strep-tagged FocA specifically retrieved PflB from Eschericha coli crude extracts. Using a bacterial two-hybrid system it could be shown that the N-terminus of FocA and the central domain of PflB were involved in the interaction. This finding was confirmed by chemical cross-linking experiments. Using constraints imposed by the amino acid residues identified in the cross-linking study we provide for the first time a model for the FocA-PflB complex. The model suggests that the N-terminus of FocA is important for interaction with PflB. An in vivo assay developed to monitor changes in formate levels in the cytoplasm revealed the importance of the interaction with PflB for optimal tranlocation of formate by FocA. This system represents a paradigm for the control of activity of FNT channel proteins.
    Graphical abstract




    Categories: Journal Articles
  • The F420-reducing [NiFe]-hydrogenase complex from Methanothermobacter marburgensis, the first X-ray structure of a group 3 family member
    [Jun 2014]

    Publication date: Available online 1 June 2014
    Source:Journal of Molecular Biology

    Author(s): Stella Vitt , Kesen Ma , Eberhard Warkentin , Johanna Moll , Antonio Pierik , Seigo Shima , Ulrich Ermler

    The reversible redox reaction between coenzyme F420 and H2 to F420H2 is catalyzed by a F420-reducing [NiFe]-hydrogenase (FrhABG) which is an enzyme of the energy metabolism of methanogenic archaea. FrhABG is a group 3 [NiFe]-hydrogenase with a dodecameric quaternary structure of 1.25 MDa as recently revealed by high resolution cryo electron microscopy. We report on the crystal structure of FrhABG from Methanothermobacter marburgensis at 1.7Å resolution and compare it with the structures of group 1 [NiFe]-hydrogenases, the only group structurally characterized yet. FrhA is similar to the large subunit of group 1 [NiFe]-hydrogenases regarding its core structure and the embedded [NiFe]-center but different because of the truncation of ca. 160 residues which results in similar but modified H2- and proton- transport pathways and in suitable interfaces for oligomerization. The small subunit FrhG is composed of a N-terminal domain related to group 1 enzymes and a new C-terminal ferredoxin-like domain carrying the distal and medial [4Fe-4S] clusters. FrhB adopts a novel fold, binds one [4Fe-4S] cluster as well as one FAD in a U-shaped conformation and provides the F420-binding site at the Si-face of the isoalloxazine ring. Similar electrochemical potentials of both catalytic reactions and the electron-transferring [4Fe-4S] clusters, determined to be E°’ ≈ –400mV, are in full agreement with the equalized forward and backward rates of the FrhABG reaction. The protein might contribute to balanced redox potentials by the aspartate coordination of the proximal [4Fe-4S] cluster, the new ferredoxin module and a rather negatively charged FAD surrounding.
    Graphical abstract




    Categories: Journal Articles
  • Editorial Board
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11









    Categories: Journal Articles
  • Contents List
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11









    Categories: Journal Articles
  • Separate Molecular Determinants in Amyloidogenic and Antimicrobial Peptides
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11

    Author(s): Michael Landreh , Jan Johansson , Hans Jörnvall

    Several amyloid-forming and antimicrobial peptides (AMYs and AMPs) have the ability to bind to and damage cell membranes. In addition, some AMYs possess antimicrobial activity and some AMPs form amyloid-like fibrils, relating the two peptide types and their properties. However, a comparison of their sequence characteristics reveals important differences. The high β-strand and aggregation propensities typical of AMYs are largely absent in α-helix-forming AMPs, which are instead marked by a strong amphipathic moment not generally found in AMYs. Although a few peptides, for example, islet amyloid polypeptide and dermaseptin S9, combine some determinants of both groups, the structural distinctions suggest that antimicrobial activity and amyloid formation are separate features not generally associated.
    Graphical abstract




    Categories: Journal Articles
  • Identification of the Active Sites in the Methyltransferases of a Transcribing dsRNA Virus
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11

    Author(s): Bin Zhu , Chongwen Yang , Hongrong Liu , Lingpeng Cheng , Feng Song , Songjun Zeng , Xiaojun Huang , Gang Ji , Ping Zhu

    Many double-stranded RNA (dsRNA) viruses are capable of transcribing and capping RNA within a stable icosahedral viral capsid. The turret of turreted dsRNA viruses belonging to the family Reoviridae is formed by five copies of the turret protein, which contains domains with both 7-N-methyltransferase and 2′-O-methyltransferase activities, and serves to catalyze the methylation reactions during RNA capping. Cypovirus of the family Reoviridae provides a good model system for studying the methylation reactions in dsRNA viruses. Here, we present the structure of a transcribing cypovirus to a resolution of ~3.8Å by cryo-electron microscopy. The binding sites for both S-adenosyl-l-methionine and RNA in the two methyltransferases of the turret were identified. Structural analysis of the turret in complex with RNA revealed a pathway through which the RNA molecule reaches the active sites of the two methyltransferases before it is released into the cytoplasm. The pathway shows that RNA capping reactions occur in the active sites of different turret protein monomers, suggesting that RNA capping requires concerted efforts by at least three turret protein monomers. Thus, the turret structure provides novel insights into the precise mechanisms of RNA methylation.
    Graphical abstract




    Categories: Journal Articles
  • Post-Translational Regulation of CD133 by ATase1/ATase2-Mediated Lysine Acetylation
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11

    Author(s): Anthony B. Mak , Mariana Pehar , Allison M.L. Nixon , Rashida A. Williams , Andrea C. Uetrecht , Luigi Puglielli , Jason Moffat

    The CD133 cell-surface protein expresses the AC133 epitope that is associated with cancer progenitor cells and cancer resistance to traditional anticancer therapies. We report that the endoplasmic reticulum Golgi intermediate compartment residing acetyltransferases, ATase1 (NAT8B) and ATase2 (NAT8), can physically interact with CD133 to acetylate the protein on three lysine residues predicted to reside on the first extracellular loop of CD133. Site-directed mutagenesis of these residues mimicking a loss of acetylation and downregulation or inhibition of ATase1/ATase2 resulted in near-complete abolishment of CD133 protein expression. We also demonstrate that targeting ATase1/ATase2 results in apoptosis of CD133 expressing acute lymphoblastic leukemia cells. Taken together, we suggest that lysine acetylation on predicted extracellular residues plays a key role in expression and trafficking of CD133 protein to the cell surface and can be targeted to disrupt CD133 regulation and function.
    Graphical abstract




    Categories: Journal Articles
  • APE1 Incision Activity at Abasic Sites in Tandem Repeat Sequences
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11

    Author(s): Mengxia Li , Jens Völker , Kenneth J. Breslauer , David M. Wilson III

    Repetitive DNA sequences, such as those present in microsatellites and minisatellites, telomeres, and trinucleotide repeats (linked to fragile X syndrome, Huntington disease, etc.), account for nearly 30% of the human genome. These domains exhibit enhanced susceptibility to oxidative attack to yield base modifications, strand breaks, and abasic sites; have a propensity to adopt non-canonical DNA forms modulated by the positions of the lesions; and, when not properly processed, can contribute to genome instability that underlies aging and disease development. Knowledge on the repair efficiencies of DNA damage within such repetitive sequences is therefore crucial for understanding the impact of such domains on genomic integrity. In the present study, using strategically designed oligonucleotide substrates, we determined the ability of human apurinic/apyrimidinic endonuclease 1 (APE1) to cleave at apurinic/apyrimidinic (AP) sites in a collection of tandem DNA repeat landscapes involving telomeric and CAG/CTG repeat sequences. Our studies reveal the differential influence of domain sequence, conformation, and AP site location/relative positioning on the efficiency of APE1 binding and strand incision. Intriguingly, our data demonstrate that APE1 endonuclease efficiency correlates with the thermodynamic stability of the DNA substrate. We discuss how these results have both predictive and mechanistic consequences for understanding the success and failure of repair protein activity associated with such oxidatively sensitive, conformationally plastic/dynamic repetitive DNA domains.
    Graphical abstract




    Categories: Journal Articles
  • Mitochondrial DNA Variant in COX1 Subunit Significantly Alters Energy Metabolism of Geographically Divergent Wild Isolates in Caenorhabditis elegans
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11

    Author(s): Stephen D. Dingley , Erzsebet Polyak , Julian Ostrovsky , Satish Srinivasan , Icksoo Lee , Amy B. Rosenfeld , Mai Tsukikawa , Rui Xiao , Mary A. Selak , Joshua J. Coon , Alexander S. Hebert , Paul A. Grimsrud , Young Joon Kwon , David J. Pagliarini , Xiaowu Gai , Theodore G. Schurr , Maik Hüttemann , Eiko Nakamaru-Ogiso , Marni J. Falk

    Mitochondrial DNA (mtDNA) sequence variation can influence the penetrance of complex diseases and climatic adaptation. While studies in geographically defined human populations suggest that mtDNA mutations become fixed when they have conferred metabolic capabilities optimally suited for a specific environment, it has been challenging to definitively assign adaptive functions to specific mtDNA sequence variants in mammals. We investigated whether mtDNA genome variation functionally influences Caenorhabditis elegans wild isolates of distinct mtDNA lineages and geographic origins. We found that, relative to N2 (England) wild-type nematodes, CB4856 wild isolates from a warmer native climate (Hawaii) had a unique p.A12S amino acid substitution in the mtDNA-encoded COX1 core catalytic subunit of mitochondrial complex IV (CIV). Relative to N2, CB4856 worms grown at 20°C had significantly increased CIV enzyme activity, mitochondrial matrix oxidant burden, and sensitivity to oxidative stress but had significantly reduced lifespan and mitochondrial membrane potential. Interestingly, mitochondrial membrane potential was significantly increased in CB4856 grown at its native temperature of 25°C. A transmitochondrial cybrid worm strain, chpIR (M, CB4856>N2), was bred as homoplasmic for the CB4856 mtDNA genome in the N2 nuclear background. The cybrid strain also displayed significantly increased CIV activity, demonstrating that this difference results from the mtDNA-encoded p.A12S variant. However, chpIR (M, CB4856>N2) worms had significantly reduced median and maximal lifespan relative to CB4856, which may relate to their nuclear–mtDNA genome mismatch. Overall, these data suggest that C. elegans wild isolates of varying geographic origins may adapt to environmental challenges through mtDNA variation to modulate critical aspects of mitochondrial energy metabolism.
    Graphical abstract




    Categories: Journal Articles
  • Assembly of Robust Bacterial Microcompartment Shells Using Building Blocks from an Organelle of Unknown Function
    [Jun 2014]

    Publication date: 29 May 2014
    Source:Journal of Molecular Biology, Volume 426, Issue 11

    Author(s): Jonathan K. Lassila , Susan L. Bernstein , James N. Kinney , Seth D. Axen , Cheryl A. Kerfeld

    Bacterial microcompartments (BMCs) sequester enzymes from the cytoplasmic environment by encapsulation inside a selectively permeable protein shell. Bioinformatic analyses indicate that many bacteria encode BMC clusters of unknown function and with diverse combinations of shell proteins. The genome of the halophilic myxobacterium Haliangium ochraceum encodes one of the most atypical sets of shell proteins in terms of composition and primary structure. We found that microcompartment shells could be purified in high yield when all seven H. ochraceum BMC shell genes were expressed from a synthetic operon in Escherichia coli. These shells differ substantially from previously isolated shell systems in that they are considerably smaller and more homogeneous, with measured diameters of 39±2nm. The size and nearly uniform geometry allowed the development of a structural model for the shells composed of 260 hexagonal units and 13 hexagons per icosahedral face. We found that new proteins could be recruited to the shells by fusion to a predicted targeting peptide sequence, setting the stage for the use of these remarkably homogeneous shells for applications such as three-dimensional scaffolding and the construction of synthetic BMCs. Our results demonstrate the value of selecting from the diversity of BMC shell building blocks found in genomic sequence data for the construction of novel compartments.
    Graphical abstract




    Categories: Journal Articles