Structural Studies of AAV2 Rep68 Reveal a Partially Structured Linker and Compact Domain Conformation.

Abstract:

:Adeno-associated virus (AAV) nonstructural proteins Rep78 and Rep68 carry out all DNA transactions that regulate the AAV life cycle. They share two multifunctional domains: an N-terminal origin binding/nicking domain (OBD) from the HUH superfamily and a SF3 helicase domain. A short linker of ∼20 amino acids that is critical for oligomerization and function connects the two domains. Although X-ray structures of the AAV5 OBD and AAV2 helicase domains have been determined, information about the full-length protein and linker conformation is not known. This article presents the solution structure of AAV2 Rep68 using small-angle X-ray scattering (SAXS). We first determined the X-ray structures of the minimal AAV2 Rep68 OBD and of the OBD with the linker region. These X-ray structures reveal novel features that include a long C-terminal α-helix that protrudes from the core of the protein at a 45° angle and a partially structured linker. SAXS studies corroborate that the linker is not extended, and we show that a proline residue in the linker is critical for Rep68 oligomerization and function. SAXS-based rigid-body modeling of Rep68 confirms these observations, showing a compact arrangement of the two domains in which they acquire a conformation that positions key residues in all domains on one face of the protein, poised to interact with DNA.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Musayev FN,Zarate-Perez F,Bardelli M,Bishop C,Saniev EF,Linden RM,Henckaerts E,Escalante CR

doi

10.1021/acs.biochem.5b00610

subject

Has Abstract

pub_date

2015-09-29 00:00:00

pages

5907-19

issue

38

eissn

0006-2960

issn

1520-4995

journal_volume

54

pub_type

杂志文章
  • Size of acetylcholine receptors in the membrane. An improved version of the radiation inactivation method.

    abstract::The radiation inactivation method was used to study the size of acetylcholine receptors in the intact membrane-bound state. This technique was reinvestigated, and modifications were made which remove substantial difficulties affecting previous applications of it to such proteins. The molecular size was deduced here by...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00538a033

    authors: Lo MM,Barnard EA,Dolly JO

    更新日期:1982-04-27 00:00:00

  • Residues 207, 216, and 221 and the catalytic activity of mGSTA1-1 and mGSTA2-2 toward benzo[a]pyrene-(7R,8S)-diol-(9S,10R)-epoxide.

    abstract::Murine class alpha glutathione S-transferase subunit types A2 (mGSTA2-2) and A1 (mGSTA1-1) have high catalytic efficiency for glutathione (GSH) conjugation of the ultimate carcinogenic metabolite of benzo[a]pyrene, (+)-anti-7,8-dihydroxy-9,10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene, [(+)-anti-BPDE]. Only 10 residues dif...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi026778+

    authors: Gu Y,Xiao B,Wargo HL,Bucher MH,Singh SV,Ji X

    更新日期:2003-02-04 00:00:00

  • Acid-base properties of ionophore A23187 in methanol-water solutions and bound to unilamellar vesicles of dimyristoylphosphatidylcholine.

    abstract::The acid-base properties of ionophore A23187 in methanol-water solutions (0--95% w/w) and bound to unilamellar vesicles of dimyristoylphosphatidylcholine were examined by ultraviolet and fluorescence spectroscopy, and the spectral properties for the acidic and basic forms were defined under these conditions. Standard ...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00539a023

    authors: Kauffman RF,Taylor RW,Pfeiffer DR

    更新日期:1982-05-11 00:00:00

  • Thermodynamic characterization of nucleoplasmin unfolding: interplay between function and stability.

    abstract::The unfolding equilibrium of recombinant (rNP) and natural variants of nucleoplasmin (NP) from Xenopus laevis has been analyzed using biochemical and spectroscopic techniques. In the presence of denaturing concentrations of guanidinium salts (GuHCl and GuSCN), both domains, core and tail, of the rNP pentamer unfold as...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi8002555

    authors: Franco G,Bañuelos S,Falces J,Muga A,Urbaneja MA

    更新日期:2008-07-29 00:00:00

  • Characterization of two glycolipid: alpha 2-3sialyltransferases, SAT-3 (CMP-NeuAc:nLcOse4Cer alpha 2-3sialyltransferase) and SAT-4 (CMP-NeuAc:GgOse4Cer alpha 2-3sialyltransferase), from human colon carcinoma (Colo 205) cell line.

    abstract::Sialyltransferase activities, SAT-3 (CMP-NeuAc:nLcOse4Cer alpha 2-3sialyltransferase) and SAT-4 (CMP-NeuAc:GgOse4Cer alpha 2-3sialyltransferase), in Colo 205 cells catalyze the transfer of sialic acid to the terminal galactose of GlcNc-- and GalNAc-containing glycolipid substrates, respectively. Competition kinetic st...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi960239l

    authors: Basu SS,Basu M,Li Z,Basu S

    更新日期:1996-04-23 00:00:00

  • A cluster of aromatic residues in the sixth membrane-spanning segment of the dopamine D2 receptor is accessible in the binding-site crevice.

    abstract::The binding site of the dopamine D2 receptor, like that of other homologous G protein-coupled receptors, is contained within a water-accessible crevice formed among its seven membrane-spanning segments. Using the substituted-cysteine accessibility method, we previously mapped the residues in the third, fifth, and seve...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi972241y

    authors: Javitch JA,Ballesteros JA,Weinstein H,Chen J

    更新日期:1998-01-27 00:00:00

  • Organization of the human protein S genes.

    abstract::Human genomic clones that span the entire protein S expressed gene (PS alpha) and the 3' two-thirds of the protein S pseudogene (PS beta) have been isolated and characterized. The PS alpha gene is greater than 80 kilobases in length and contains 14 introns and 15 exons, as well as 6 repetitive "Alu" sequences. Exons I...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00486a010

    authors: Schmidel DK,Tatro AV,Phelps LG,Tomczak JA,Long GL

    更新日期:1990-08-28 00:00:00

  • Engineering the independent folding of the subtilisin BPN' pro-domain: correlation of pro-domain stability with the rate of subtilisin folding.

    abstract::The 77-amino acid pro-domain greatly accelerates the in vitro folding of subtilisin in a bimolecular reaction whose product is a tight complex between folded subtilisin and folded pro-domain. In this complex the pro-domain has a compact structure with a four-stranded antiparallel beta-sheet and two three-turn alpha-he...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi972741r

    authors: Wang L,Ruan B,Ruvinov S,Bryan PN

    更新日期:1998-03-03 00:00:00

  • Concentration of activated intermediates of the fructose-1,6-bisphosphate aldolase and triosephosphate isomerase reactions.

    abstract::As discovered by Grazi & Trombetta [Grazi, E., & Trombetta, G. (1978) Biochem. J. 175, 361], fructose-1,6-bisphosphate aldolase of rabbit muscle causes the slow formation of inorganic phosphate (Pi) and methylglyoxal when incubated with dihydroxyacetone phosphate (DHAP). In addition, these authors found an acid-labile...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00508a026

    authors: Iyengar R,Rose IA

    更新日期:1981-03-03 00:00:00

  • Diversity and abundance of polyadenylated RNA from Achlya ambisexualis.

    abstract::The diversity, abundance, and DNA sequence representation of poly(adenylic acid) containing RNA derived from cells of Achlya ambisexualis cultured in defined and undefined media have been determined. The kinetics of hybridization of polyadenylated RNA with complementary DNA were the same for both culture conditions an...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00597a025

    authors: Rozek CE,Orr WC,Timberlake WE

    更新日期:1978-02-21 00:00:00

  • A second photochromic bacteriophytochrome from Synechocystis sp. PCC 6803: spectral analysis and down-regulation by light.

    abstract::It now appears that photosynthetic prokaryotes and lower eukaryotes possess higher plant phytochrome-like proteins. In this work, a second phytochrome-like gene was isolated, in addition to the recently identified Cph1 phytochrome, from the Synechocystis sp. PCC 6803, and its gene product was characterized photochemic...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi992831r

    authors: Park CM,Kim JI,Yang SS,Kang JG,Kang JH,Shim JY,Chung YH,Park YM,Song PS

    更新日期:2000-09-05 00:00:00

  • Carboxylases in de novo purine biosynthesis. Characterization of the Gallus gallus bifunctional enzyme.

    abstract::Two successive steps in de novo purine biosynthesis are catalyzed by the enzymes 5-aminoimidazole ribonucleotide (AIR) carboxylase and 4-[(N-succinylamino)carbonyl]-5-aminoimidazole ribonucleotide (SAICAR) synthetase. Amino acid sequence alignments of the proteins from various sources suggested that several unusual di...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00205a030

    authors: Firestine SM,Davisson VJ

    更新日期:1994-10-04 00:00:00

  • Fourier transform infrared spectroscopy and electrochemistry of the primary electron donor in Rhodobacter sphaeroides and Rhodopseudomonas viridis reaction centers: vibrational modes of the pigments in situ and evidence for protein and water modes affecte

    abstract::Protein electrochemistry in an ultra-thin-layer electrochemical cell suitable for UV/vis and IR spectroscopy has been used to characterize the vibrational modes of the primary electron donors of Rhodobacter sphaeroides and Rhodopseudomonas viridis reaction centers in their neutral and cation radical states (P and P+, ...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00068a007

    authors: Leonhard M,Mäntele W

    更新日期:1993-05-04 00:00:00

  • Localization of a reactive exofacial sulfhydryl on the glucose carrier of human erythrocytes.

    abstract::Tryptic digestion studies of the human erythrocyte glucose carrier have shown that a reactive and transport-sensitive exofacial sulfhydryl is located in the carboxy-terminal half of the molecule, corresponding to Cys347, Cys421, or Cys429. In the present studies, the erythrocyte glucose carrier labeled on the exofacia...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00497a014

    authors: May JM,Buchs A,Carter-Su C

    更新日期:1990-11-13 00:00:00

  • Unfolding rates of barstar determined in native and low denaturant conditions indicate the presence of intermediates.

    abstract::The folding and unfolding rates of the small protein, barstar, have been monitored using stopped-flow measurements of intrinsic tryptophan fluorescence at 25 degrees C, pH 8.5, and have been compared over a wide range of urea and guanidine hydrochloride (GdnHCl) concentrations. When the logarithms of the rates of fold...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi011494v

    authors: Sridevi K,Udgaonkar JB

    更新日期:2002-02-05 00:00:00

  • NMR structure and molecular dynamics of the in-plane membrane anchor of nonstructural protein 5A from bovine viral diarrhea virus.

    abstract::Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) is a monotopic membrane protein anchored to the membrane by an N-terminal in-plane amphipathic alpha-helix. This membrane anchor is essential for the assembly of a functional viral replication complex. Although amino acid sequences differ considerably, putative m...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi0517685

    authors: Sapay N,Montserret R,Chipot C,Brass V,Moradpour D,Deléage G,Penin F

    更新日期:2006-02-21 00:00:00

  • An appraisal of multiple NADPH binding-site models proposed for cytochrome P450 reductase, NO synthase, and related diflavin reductase systems.

    abstract::The diflavin reductases exemplified by mammalian cytochrome P450 reductase catalyze NADPH dehydrogenation and electron transfer to an associated monooxygenase. It has recently been proposed that double occupancy of the NADPH dehydrogenation site inhibits the NADPH to FAD hydride transfer step in this series of enzymes...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi0360408

    authors: Daff S

    更新日期:2004-04-06 00:00:00

  • Cembranoid and long-chain alkanol sites on the nicotinic acetylcholine receptor and their allosteric interaction.

    abstract::Long-chain alkanols are general anesthetics which can also act as uncharged noncompetitive inhibitors of the peripheral nicotinic acetylcholine receptor (AChR) by binding to one or more specific sites on the AChR. Cembranoids are naturally occurring, uncharged noncompetitive inhibitors of peripheral and neuronal AChRs...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi0112255

    authors: Pagán OR,Eterović VA,Garcia M,Vergne D,Basilio CM,Rodríguez AD,Hann RM

    更新日期:2001-09-18 00:00:00

  • Large activation energy barriers to chaperone-peptide complex formation and dissociation.

    abstract::To probe the mechanism of chaperone substrate selection, we have investigated the kinetics of complex formation and dissociation between the molecular chaperone DnaK and a short peptide (Cro, representing amino acids 1-12 of the cro repressor protein). The Cro protein was N-terminally labeled with the environmentally ...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00047a024

    authors: Farr CD,Galiano FJ,Witt SN

    更新日期:1995-11-28 00:00:00

  • Effect of thioredoxin deletion and p53 cysteine replacement on human p53 activity in wild-type and thioredoxin reductase null yeast.

    abstract::Reporter gene transactivation by human p53 is inhibited in budding yeast lacking the TRR1 gene encoding thioredoxin reductase. To investigate the role of thioredoxin in controlling p53 activity, the level of reporter gene transactivation by p53 was determined in yeast lacking the TRX1 and TRX2 genes encoding cytosolic...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi900757q

    authors: Stoner CS,Pearson GD,Koç A,Merwin JR,Lopez NI,Merrill GF

    更新日期:2009-09-29 00:00:00

  • Mechanism of the interaction of beta(2)-glycoprotein I with negatively charged phospholipid membranes.

    abstract::In an attempt to understand the multifunctional involvement of beta(2)-glycoprotein I (beta(2)GPI) in autoimmune diseases, thrombosis, atherosclerosis, and inflammatory processes, substantial interest is focused on the interaction of beta(2)GPI with negatively charged ligands, in particular, with acidic phospholipids....

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi0114372

    authors: Hammel M,Schwarzenbacher R,Gries A,Kostner GM,Laggner P,Prassl R

    更新日期:2001-11-27 00:00:00

  • Posttranslational modifications of the Ia-associated invariant protein p41 after gene transfer.

    abstract::Biochemical analysis of a rat fibroblast cell clone, transfected with the murine Ia associated invariant chain gene, demonstrates the expression of a family of proteins. This indicates that all members of the invariant protein family are derived from the same gene. The proteins p41, Ii, p27, p25, and p10, synthesized ...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00411a028

    authors: Koch N

    更新日期:1988-05-31 00:00:00

  • Crystal structure of Thermobifida fusca endoglucanase Cel6A in complex with substrate and inhibitor: the role of tyrosine Y73 in substrate ring distortion.

    abstract::Endoglucanase Cel6A from Thermobifida fusca hydrolyzes the beta-1,4 linkages in cellulose at accessible points along the polymer. The structure of the catalytic domain of Cel6A from T. fusca in complex with a nonhydrolysable substrate analogue that acts as an inhibitor, methylcellobiosyl-4-thio-beta-cellobioside (Glc(...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi0506730

    authors: Larsson AM,Bergfors T,Dultz E,Irwin DC,Roos A,Driguez H,Wilson DB,Jones TA

    更新日期:2005-10-04 00:00:00

  • Substrate specificity and excision kinetics of Escherichia coli endonuclease VIII (Nei) for modified bases in DNA damaged by free radicals.

    abstract::Endonuclease VIII (Nei) is one of three enzymes in Escherichia coli that are involved in base-excision repair of oxidative damage to DNA. We investigated the substrate specificity and excision kinetics of this DNA glycosylase for bases in DNA that have been damaged by free radicals. Two different DNA substrates were p...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi015552o

    authors: Dizdaroglu M,Burgess SM,Jaruga P,Hazra TK,Rodriguez H,Lloyd RS

    更新日期:2001-10-09 00:00:00

  • The amino- and carboxyl-terminal fragments of the Bacillus thuringensis Cyt1Aa toxin have differential roles in toxin oligomerization and pore formation.

    abstract::The Cyt toxins produced by the bacteria Bacillus thuringiensis show insecticidal activity against some insects, mainly dipteran larvae, being able to kill mosquitoes and black flies. However, they also possess a general cytolytic activity in vitro, showing hemolytic activity in red blood cells. These proteins are comp...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi101239r

    authors: Rodriguez-Almazan C,Ruiz de Escudero I,Cantón PE,Muñoz-Garay C,Pérez C,Gill SS,Soberón M,Bravo A

    更新日期:2011-01-25 00:00:00

  • Raman difference spectroscopic studies of the myosin S1.MgADP.vanadate complex.

    abstract::The Raman spectra of the nonbridging V--O bonds in the myosin S1.MgADP.Vi complex, often believed to be a transition-state analogue for the phosphotransfer reaction catalyzed by myosin, and in a vanadate solution model compound have been obtained using Raman difference spectroscopic techniques. A symmetric/asymmetric ...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi980556n

    authors: Deng H,Wang J,Callender RH,Grammer JC,Yount RG

    更新日期:1998-08-04 00:00:00

  • The room temperature reaction of carbon monoxide and oxygen with the cytochrome bd quinol oxidase from Escherichia coli.

    abstract::When grown under O2-limited conditions, Escherichia coli expresses a cytochrome bd quinol oxidase that has an unusually high affinity for O2. We have studied the reaction of cytochrome bd with CO and O2 by rapid-reaction spectrophotometry. The reduced enzyme forms a photosensitive ferrocytochrome d-CO complex, and fol...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00254a021

    authors: Hill BC,Hill JJ,Gennis RB

    更新日期:1994-12-20 00:00:00

  • Glycosphingolipid backbone conformation and behavior in cholesterol-containing phospholipid bilayers.

    abstract::2H NMR spectroscopy was used to consider correspondence between existing single-crystal X-ray data for glycosphingolipids and their ceramide backbone conformation in fluid phospholipid membranes. A monoglycosylated sphingolipid, glucosylceramide (GlcCer), which represents the core structure of many important glycosphi...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00066a024

    authors: Hamilton KS,Jarrell HC,Brière KM,Grant CW

    更新日期:1993-04-20 00:00:00

  • Isolation and characterization of titin T1 from bovine cardiac muscle.

    abstract::Methods were developed for isolation of bovine cardiac titin in the apparently full-sized T1 form. The pure protein could be obtained either in the denatured state after chromatography of myofibril extracts on Sephacryl S-1000 or in the native state using hydroxylapatite chromatography. Titin-specific chicken polyclon...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/bi00193a012

    authors: Pan KM,Damodaran S,Greaser ML

    更新日期:1994-07-12 00:00:00

  • Substrate-Induced Facilitated Dissociation of the Competitive Inhibitor from the Active Site of O-Acetyl Serine Sulfhydrylase Reveals a Competitive-Allostery Mechanism.

    abstract::By classical competitive antagonism, a substrate and competitive inhibitor must bind mutually exclusively to the active site. The competitive inhibition of O-acetyl serine sulfhydrylase (OASS) by the C-terminus of serine acetyltransferase (SAT) presents a paradox, because the C-terminus of SAT binds to the active site...

    journal_title:Biochemistry

    pub_type: 杂志文章

    doi:10.1021/acs.biochem.7b00500

    authors: Singh AK,Ekka MK,Kaushik A,Pandya V,Singh RP,Banerjee S,Mittal M,Singh V,Kumaran S

    更新日期:2017-09-19 00:00:00