Engineering activity and stability of Thermotoga maritima glutamate dehydrogenase. II: construction of a 16-residue ion-pair network at the subunit interface.

Abstract:

:The role of an 18-residue ion-pair network, that is present in the glutamate dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus, in conferring stability to other, less stable homologous enzymes, has been studied by introducing four new charged amino acid residues into the subunit interface of glutamate dehydrogenase from the hyperthermophilic bacterium Thermotoga maritima. These two GDHs are 55 % identical in amino acid sequence, differ greatly in thermo-activity and stability and derive from microbes with different phylogenetic positions. Amino acid substitutions were introduced as single mutations as well as in several combinations. Elucidation of the crystal structure of the quadruple mutant S128R/T158E/N117R/S160E T. maritima glutamate dehydrogenase showed that all anticipated ion-pairs are formed and that a 16-residue ion-pair network is present. Enlargement of existing networks by single amino acid substitutions unexpectedly resulted in a decrease in resistance towards thermal inactivation and thermal denaturation. However, combination of destabilizing single mutations in most cases restored stability, indicating the need for balanced charges at subunit interfaces and high cooperativity between the different members of the network. Combination of the three destabilizing mutations in triple mutant S128R/T158E/N117R resulted in an enzyme with a 30 minutes longer half-life of inactivation at 85 degrees C, a 3 degrees C higher temperature optimum for catalysis, and a 0.5 degrees C higher apparent melting temperature than that of wild-type glutamate dehydrogenase. These findings confirm the hypothesis that large ion-pair networks do indeed stabilize enzymes from hyperthermophilic organisms.

journal_name

J Mol Biol

authors

Lebbink JH,Knapp S,van der Oost J,Rice D,Ladenstein R,de Vos WM

doi

10.1006/jmbi.1999.2779

keywords:

subject

Has Abstract

pub_date

1999-06-04 00:00:00

pages

357-69

issue

2

eissn

0022-2836

issn

1089-8638

pii

S0022-2836(99)92779-1

journal_volume

289

pub_type

杂志文章
  • The innermost chorionic layer of Drosophila. I. The role of chorin octamers in the formation of a family of interdigitating crystalline plates.

    abstract::The innermost chorionic layer (ICL) within egg shells of Drosophila melanogaster is composed of thin, abutting three-dimensional crystalline plates which form a closed, membrane-like sheath. Collectively, the crystals within the sheath appear to form a family of related three-dimensional crystals in space group C222; ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(87)90350-0

    authors: Akey CW,Edelstein SJ

    更新日期:1987-02-20 00:00:00

  • Increased thermostability and fidelity of DNA synthesis of wild-type and mutant HIV-1 group O reverse transcriptases.

    abstract::Reverse transcription coupled with DNA amplification has become a well-established and powerful molecular technique for studying ribonucleic acids. However, the efficiency of those reactions is largely dependent on the molecular properties of currently used reverse transcriptases (RTs). Engineered and natural RT varia...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2009.07.081

    authors: Alvarez M,Matamoros T,Menéndez-Arias L

    更新日期:2009-10-02 00:00:00

  • The Spliceosomal Protein SF3B5 is a Novel Component of Drosophila SAGA that Functions in Gene Expression Independent of Splicing.

    abstract::The interaction between splicing factors and the transcriptional machinery provides an intriguing link between the coupled processes of transcription and splicing. Here, we show that the two components of the SF3B complex, SF3B3 and SF3B5, that form part of the U2 small nuclear ribonucleoprotein particle (snRNP) are a...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2016.05.009

    authors: Stegeman R,Spreacker PJ,Swanson SK,Stephenson R,Florens L,Washburn MP,Weake VM

    更新日期:2016-09-11 00:00:00

  • Crystal structures of artocarpin, a Moraceae lectin with mannose specificity, and its complex with methyl-alpha-D-mannose: implications to the generation of carbohydrate specificity.

    abstract::The seeds of jack fruit (Artocarpus integrifolia) contain two tetrameric lectins, jacalin and artocarpin. Jacalin was the first lectin found to exhibit the beta-prism I fold, which is characteristic of the Moraceae plant lectin family. Jacalin contains two polypeptide chains produced by a post-translational proteolysi...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.2001.5432

    authors: Pratap JV,Jeyaprakash AA,Rani PG,Sekar K,Surolia A,Vijayan M

    更新日期:2002-03-22 00:00:00

  • Empirical statistical estimates for sequence similarity searches.

    abstract::The FASTA package of sequence comparison programs has been modified to provide accurate statistical estimates for local sequence similarity scores with gaps. These estimates are derived using the extreme value distribution from the mean and variance of the local similarity scores of unrelated sequences after the score...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1997.1525

    authors: Pearson WR

    更新日期:1998-02-13 00:00:00

  • X-ray structure of the quinoprotein ethanol dehydrogenase from Pseudomonas aeruginosa: basis of substrate specificity.

    abstract::The homodimeric enzyme form of quinoprotein ethanol dehydrogenase from Pseudomonas aeruginosa ATCC 17933 crystallizes readily with the space group R3. The X-ray structure was solved at 2.6 A resolution by molecular replacement. Aside from differences in some loops, the folding of the enzyme is very similar to the larg...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.2000.3603

    authors: Keitel T,Diehl A,Knaute T,Stezowski JJ,Höhne W,Görisch H

    更新日期:2000-04-07 00:00:00

  • Purification and secondary structure determination of simian immunodeficiency virus p27.

    abstract::We have developed a novel method for the expression and purification of p27, the major core protein of simian immunodeficiency virus. Circular dichroism measurements of purified p27 were used to determine the relative amounts of alpha-helix, beta-sheet and unordered secondary structural elements. These empirically det...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/s0022-2836(05)80308-0

    authors: Burns NR,Craig S,Lee SR,Richardsons MH,Stenner N,Adams SE,Kingsman SM,Kingsman AJ

    更新日期:1990-11-20 00:00:00

  • Vibrio cholerae YaeO is a Structural Homologue of RNA Chaperone Hfq that Inhibits Rho-dependent Transcription Termination by Dissociating its Hexameric State.

    abstract::Rho-dependent transcription termination is a well-conserved process in bacteria. The Psu and YaeO proteins are the two established inhibitors of the ATP-dependent RNA helicase Rho protein of Escherichia coli. Here, we show a detailed sequence and phylogenetic analysis demonstrating that Vibrio cholerae YaeO (VcYaeO) i...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2019.09.019

    authors: Pal K,Yadav M,Jain S,Ghosh B,Sen R,Sen U

    更新日期:2019-12-06 00:00:00

  • The salt-dependence of a protein-ligand interaction: ion-protein binding energetics.

    abstract::Using the binding of a nucleotide inhibitor (guanosine-3'-monophosphate) to a ribonuclease (ribonuclease Sa) as a model system, we show that the salt-dependence of the interaction arises due to specific ion binding at the site of nucleotide binding. The presence of specific ion-protein binding is concluded from a comb...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2004.12.018

    authors: Waldron TT,Schrift GL,Murphy KP

    更新日期:2005-02-25 00:00:00

  • Shaping the Immune Landscape in Cancer by Galectin-Driven Regulatory Pathways.

    abstract::Along with the discovery of tumor-driven inflammatory pathways, there has been a considerable progress over the past 10years in understanding the mechanisms leading to cancer immunosurveillance and immunoediting. Several regulatory pathways, typically involved in immune cell homeostasis, are co-opted by cancer cells t...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章,评审

    doi:10.1016/j.jmb.2016.03.021

    authors: Rabinovich GA,Conejo-García JR

    更新日期:2016-08-14 00:00:00

  • Position-specific inhibition of yeast mitochondrial transcription by a poly(T) sequence.

    abstract::The 3' flanking nucleotide(s) of the octanucleotide promoter sequence regulates transcriptional efficiency of some mitochondrial genes in Saccharomyces cerevisiae. To understand this regulation the in vitro transcriptional activity of various synthetic mitochondrial promoters carrying different 3' flanking sequences w...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1997.1483

    authors: Biswas TK,Getz GS

    更新日期:1998-01-30 00:00:00

  • Allosteric control by ATP of non-folded protein binding to GroEL.

    abstract::Co-operativity in ATP hydrolysis by GroEL can be described by a model in which each ring of GroEL is in equilibrium between a low (T) and high (R) affinity state for ATP. According to this model, the GroEL double-ring is in equilibrium between three states: TT, TR and RR. In order to find out which states bind non-fol...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1996.0028

    authors: Yifrach O,Horovitz A

    更新日期:1996-01-26 00:00:00

  • Crystal structures of an intein from the split dnaE gene of Synechocystis sp. PCC6803 reveal the catalytic model without the penultimate histidine and the mechanism of zinc ion inhibition of protein splicing.

    abstract::The first naturally occurring split intein was found in the dnaE gene of Synechocystis sp. PCC6803 and belongs to a subclass of inteins without a penultimate histidine residue. We describe two high-resolution crystal structures, one derived from an excised Ssp DnaE intein and the second from a splicing-deficient precu...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2005.09.039

    authors: Sun P,Ye S,Ferrandon S,Evans TC,Xu MQ,Rao Z

    更新日期:2005-11-11 00:00:00

  • DNA-directed DNA polymerase and strand displacement activity of the reverse transcriptase encoded by the R2 retrotransposon.

    abstract::R2 elements are non-long terminal repeat (non-LTR) retrotransposons with a single open reading-frame encoding reverse transcriptase, DNA endonuclease and nucleic acid-binding domains. The elements are specialized for insertion into the 28 S rRNA genes of many animal phyla. The R2-encoded activities initiate retrotrans...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2007.09.047

    authors: Kurzynska-Kokorniak A,Jamburuthugoda VK,Bibillo A,Eickbush TH

    更新日期:2007-11-23 00:00:00

  • Three-dimensional solid-state NMR study of a seven-helical integral membrane proton pump--structural insights.

    abstract::Proteorhodopsin (PR) is a recently discovered ubiquitous eubacterial retinal-binding light-driven proton pump. Almost 1000 PR variants are widely distributed in species of marine and freshwater bacteria, suggesting PR's important photobiological role. PR is a typical seven-transmembrane alpha-helical membrane protein ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2009.01.011

    authors: Shi L,Ahmed MA,Zhang W,Whited G,Brown LS,Ladizhansky V

    更新日期:2009-03-06 00:00:00

  • Determination of the MurD mechanism through crystallographic analysis of enzyme complexes.

    abstract::UDP -N- acetylmuramoyl- L -alanine: D -glutamate (MurD) ligase catalyses the addition of d -glutamate to the nucleotide precursor UDP -N- acetylmuramoyl- L -alanine (UMA). The crystal structures of three complexes of Escherichia coli MurD with a variety of substrates and products have been determined to high resolutio...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1999.2800

    authors: Bertrand JA,Auger G,Martin L,Fanchon E,Blanot D,Le Beller D,van Heijenoort J,Dideberg O

    更新日期:1999-06-11 00:00:00

  • Molecular and structural basis for redox regulation of beta-actin.

    abstract::An essential consequence of growth factor-mediated signal transduction is the generation of intracellular H(2)O(2). It operates as a second messenger in the control of actin microfilament dynamics, causing rapid and dramatic changes in the morphology and motile activity of stimulated cells. Little is understood about ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2007.04.056

    authors: Lassing I,Schmitzberger F,Björnstedt M,Holmgren A,Nordlund P,Schutt CE,Lindberg U

    更新日期:2007-07-06 00:00:00

  • Multiple sequence threading: an analysis of alignment quality and stability.

    abstract::Methods that compare a protein sequence directly to a structure can be divided into those that construct a molecular model (threading methods) and those that perform a sequence alignment with the structure encoded as a sequence of structural states (one-dimensional/three-dimensional (1D/3D) matching). The former take ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1997.1008

    authors: Taylor WR

    更新日期:1997-06-27 00:00:00

  • Mia40 combines thiol oxidase and disulfide isomerase activity to efficiently catalyze oxidative folding in mitochondria.

    abstract::Mia40 (a mitochondrial import and assembly protein) catalyzes disulfide bond formation in proteins in the mitochondrial intermembrane space. By using Cox17 (a mitochondrial copper-binding protein) as a natural substrate, we discovered that, in the presence of Mia40, the formation of native disulfides is strongly favor...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2014.10.022

    authors: Koch JR,Schmid FX

    更新日期:2014-12-12 00:00:00

  • On the connection between inherent DNA flexure and preferred binding of hydroxymethyluracil-containing DNA by the type II DNA-binding protein TF1.

    abstract::TF1 is a member of the family of type II DNA-binding proteins, which also includes the bacterial HU proteins and the Escherichia coli integration host factor (IHF). Distinctive to TF1, which is encoded by the Bacillus subtilis bacteriophage SPO1, is its preferential binding to DNA in which thymine is replaced by 5-hyd...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1996.0392

    authors: Grove A,Galeone A,Mayol L,Geiduschek EP

    更新日期:1996-07-12 00:00:00

  • Contributions of residue pairing to beta-sheet formation: conservation and covariation of amino acid residue pairs on antiparallel beta-strands.

    abstract::In an effort to better understand beta-sheet assembly, we have investigated the evolutionary behavior of neighboring residues on adjacent antiparallel beta-strands. Residue pairs were classified according to solvent exposure as well as by whether their backbone NH and C==O groups are hydrogen bonded. The conservation ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.2000.4364

    authors: Mandel-Gutfreund Y,Zaremba SM,Gregoret LM

    更新日期:2001-02-02 00:00:00

  • In-vivo degradation pathway of an excised intervening sequence.

    abstract::A study of the distribution in heterogeneous nuclear RNP (hnRNP) of the different forms of the first intervening sequence (IVSI) from the E3 transcription unit of adenovirus serotype 2 suggests a three-step pathway for IVS excision and degradation. First, the lariat IVS is formed in a large RNP of the size of premesse...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(87)90480-3

    authors: Sittler A,Gallinaro H,Kister L,Jacob M

    更新日期:1987-10-20 00:00:00

  • Deletion analysis of the lambda tR1 termination region. Effect of sequences near the transcript release sites, and the minimum length of rho-dependent transcripts.

    abstract::In order to determine how much of the natural sequence is required for function of the lambda tR1 rho-dependent terminator, and to determine the minimum length required, we made two deletion series of a tR1 derivative that contains mostly foreign DNA, encoding C-rich RNA, substituted for the natural upstream sequences...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1994.1229

    authors: Hart CM,Roberts JW

    更新日期:1994-04-01 00:00:00

  • Control of actin dynamics in cell motility.

    abstract::Actin polymerization plays a major role in cell movement. The controls of actin sequestration/desequestration and of filament turnover are two important features of cell motility. Actin binding proteins use properties derived from the steady-state monomer-polymer cycle of actin in the presence of ATP, to control the F...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章,评审

    doi:10.1006/jmbi.1997.1062

    authors: Carlier MF,Pantaloni D

    更新日期:1997-06-20 00:00:00

  • Monomers of the Escherichia coli SSB-1 mutant protein bind single-stranded DNA.

    abstract::The Escherichia coli wild-type single strand binding (SSB) protein is a stable tetramer that binds to single-stranded (ss) DNA in its role in DNA replication, recombination and repair. The ssb-1 mutation, a substitution of tyrosine for histidine-55 within the SSB-1 protein, destabilizes the tetramer with respect to mo...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(91)90611-9

    authors: Bujalowski W,Lohman TM

    更新日期:1991-01-05 00:00:00

  • Zooming in on Transcription Preinitiation.

    abstract::Class II gene transcription commences with the assembly of the Preinitiation Complex (PIC) from a plethora of proteins and protein assemblies in the nucleus, including the General Transcription Factors (GTFs), RNA polymerase II (RNA pol II), co-activators, co-repressors, and more. TFIID, a megadalton-sized multiprotei...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章,评审

    doi:10.1016/j.jmb.2016.04.003

    authors: Gupta K,Sari-Ak D,Haffke M,Trowitzsch S,Berger I

    更新日期:2016-06-19 00:00:00

  • Crystal structure of a human embryonic haemoglobin: the carbonmonoxy form of gower II (alpha2 epsilon2) haemoglobin at 2.9 A resolution.

    abstract::The production of recombinant embryonic haemoglobins via a yeast expression system has enabled structural and functional studies to be conducted on these proteins. As part of a programme aimed at understanding the properties of the embryonic haemoglobins we have crystallized the human alpha2 epsilon2 (Gower II) embryo...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1998.1868

    authors: Sutherland-Smith AJ,Baker HM,Hofmann OM,Brittain T,Baker EN

    更新日期:1998-07-17 00:00:00

  • Distinct Roles of Mic12 and Mic27 in the Mitochondrial Contact Site and Cristae Organizing System.

    abstract::The mitochondrial inner membrane consists of two morphologically distinct domains, the inner boundary membrane and large invaginations termed cristae. Narrow membrane structures, the crista junctions, link these two domains. Maintenance of this elaborate architecture depends on the evolutionarily conserved mitochondri...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2016.02.031

    authors: Zerbes RM,Höß P,Pfanner N,van der Laan M,Bohnert M

    更新日期:2016-04-24 00:00:00

  • Structural insight into how an anti-idiotypic antibody against D3H44 (anti-tissue factor antibody) restores normal coagulation.

    abstract::6A6 is a murine monoclonal antibody raised against the humanized anti-tissue factor antibody D3H44. 6A6 is able to completely neutralize the anticoagulant activity of D3H44 in tissue factor-dependent functional assays, such as endotoxin-induced whole blood clotting, prothrombin time, as well as factor X and factor IX ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/s0022-2836(03)00735-6

    authors: Eigenbrot C,Meng YG,Krishnamurthy R,Lipari MT,Presta L,Devaux B,Wong T,Moran P,Bullens S,Kirchhofer D

    更新日期:2003-08-08 00:00:00

  • A study of electrophoretic mobility of DNA in agarose and polyacrylamide gels.

    abstract::The aim of this paper is to clarify the mechanism of gel electrophoresis of DNA under constant-field conditions. We have conducted a large number of experiments on double-stranded DNA varying in length between approximately 10 and approximately 50,000 base-pairs, in both agarose and polyacrylamide gels ranging from 0....

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(91)80187-y

    authors: Calladine CR,Collis CM,Drew HR,Mott MR

    更新日期:1991-10-05 00:00:00