171,667 research outputs found
Delivery truck for Thomas H. Muth, automobile dealer in Paterson, New Jersey
No full textDelivery truck for Thomas H. Muth, automobile dealer, 644 Madison Avenue, circa 1915. The truck driver has a new stock of "Fisk Auto Cases and Tubes" wrapped in burlap. Sign reads on truck: Time to retire, buy Fisk auto cases and tubes, auto accessories and C
The Escherichia coli MutL Protein Physically Interacts with MutH and Stimulates the MutH-associated Endonuclease Activity
No full textAll possible pairwise combinations of UvrD, MutL, MutS, and MutH were tested using the yeast two-hybrid system to identify potential interactions involving mismatch repair proteins. A two-hybrid screen previously identified a physical interaction between MutL and UvrD. Although several other known interactions were not observed, a novel interaction between MutL and MutH was detected. A series of truncations from the NH2 and COOH termini of MutL demonstrated that the COOH-terminal 218 amino acids were sufficient for the two-hybrid interaction with MutH. Removal of a small number of residues from either the NH2 or COOH termini of MutH eliminated the two-hybrid interaction with MutL. Protein affinity chromatography experiments confirmed that MutL, but not MutS, physically associates with MutH. Furthermore, MutL greatly stimulated the d(GATC)-specific endonuclease activity of MutH in the absence of MutS and a mispaired base. Stimulation of the MutH-associated endonuclease activity by MutL was dependent on ATP binding but not ATP hydrolysis. Further stimulation of this reaction by MutS required the presence of a DNA mismatch and a hydrolyzable form of ATP. These results suggest that MutL activates the MutH-associated endonuclease activity through a physical interaction during methyl-directed mismatch repair in Escherichia coli
Mutational analysis of the MutH protein from Escherichia coli
No full textSite-directed mutagenesis was performed on several areas of MutH based on the similarity of MutH and PvuII structural models. The aims were to identify DNA-binding residues; to determine whether MutH has the same mechanism for DNA binding and catalysis as PvuII; and to localize the residues responsible for MutH stimulation by MutL. No DNA-binding residues were identified in the two flexible loop regions of MutH, although similar loops in PvuII are involved in DNA binding. Two histidines in MutH are in a similar position as two histidines (His-84 and His-85) in PvuII that signal for DNA binding and catalysis. These MutH histidines (His-112 and His-115) were changed to alanines, but the mutant proteins had wild-type activity both in vivo and in vitro. The results indicate that the MutH signal for DNA binding and catalysis remains unknown. Instead, a lysine residue (Lys-48) was found in the first flexible loop that functions in catalysis together with the three presumed catalytic amino acids (Asp-70, Glu-77, and Lys-79). Two deletion mutations (MutHDelta224 and MutHDelta214) in the C-terminal end of the protein, localized the MutL stimulation region to five amino acids (Ala-220, Leu-221, Leu-222, Ala-223, and Arg-224)
Mutational Analysis of the MutH from Escherichia Coli: a Dissertation
No full textSome images did not scan well. Please see print version for images. Page 130 missing in original document.DNA mismatch repair is one process in the preservation of genomic integrity. It has been found in Archeae, bacteria, plants, yeast and mammals. The mismatch repair system is highly conserved among species and allows the strand-specific elimination of base-base mispairs, chemical base modifications, as well as short insertion/deletion loops following DNA replication. The repair system also has important effects on homeologous recombination, contributing to the frequency of reciprocal exchanges. In humans, defects in the repair system have been found to be associated with tumorigenesis. In Escherichia coli, this pathway was originally called long patch repair before being renamed the methyl-directed mismatch repair system. It is unique in that it utilizes a DNA methylation pattern to discriminate between the parental DNA strand and the newly synthesized daughter DNA strand. The current model for the initiation of methyl-directed mismatch repair is that the mispaired bases are recognized and bound by the MutS protein with MutL as a helper protein for binding. MutL also assists the MutH protein to bind, thereby forming the completed initiation complex of MutS, MutL and MutH. In the presence of ATP, there is evidence for translocation ofthe complex along the DNA forming alpha loops. At a d(GATC) site the MutH protein binds and nicks the unmethylated daughter DNA strand 5' to the d(G) (by recognizing the N6-d(A) methylation of the parental DNA strand which it is unable to cut). This completes the initiation of the repair system and allows the hydrolysis and resynthesis of the daughter DNA strand. MutH is a monomer of 25.5 kD in solution and contains a latent Mg2+-dependent endonuclease activity. Unmethylated DNA is nicked without any discrimination on one of the two strands and fully methylated DNA is resistant to cleavage by MutH even though the protein is able to bind the d(GATC) site. The structure of MutH was recently solved and compared to a group of restriction endonucleases that share a structural common core domain with similarly placed catalytic residues. The MutH protein is comprised of two major domains that are able to pivot and rotate with respect to one another. The cleft between the two domains is large enough for double-strand DNA to bind. This research started with the determination of the MutH structure before it was known. After crystallizing the protein and collecting several heavy atom data sets, it was found that the electron density maps were too discontinuous to trace the structure of the protein. Following that work, site-directed mutagenesis was performed on several areas of MutH based on the similarity of MutH and PvuII structural models. The aims were to identify DNA binding residues (in two flexible loop regions), to determine if MutH has the same mechanism for DNA binding and catalysis as PvuII (MutH histidines 112 and 115), and to localize the residues responsible for MutH stimulation by MutL (MutH C-terminal tail region). An in-vivoscreen based on the mutator phenotype was used to select for functionally defective MutH mutants. These bacteria accumulate mutations at a greater frequency than wild-type and this was monitored by selection on plates with rifampicin. Three MutH mutants were identified from this screen (K48A, G49A, and Δ214). They were purified and assayed for total activity and binding ability. Four other mutants with wild-type phenotypic screen results were also chosen to confirm they were not involved in any MutH function (D47A, H112A, H115A, and Δ224). No DNA binding residues (such as D47A) were identified in the two flexible loop regions of MutH, although similar loops in PvuII are involved in DNA binding. The purified D47A MutH protein showed wild-type biochemical activity. Instead, the lysine residue (K48) in the first flexible loop was found to function in catalysis together with the three presumed catalytic amino acids (Asp70, Glu77, and Lys79). This purified MutH protein (K48A) had wild-type binding ability but no endonuclease activity without MutL. In the presence of MutL, the K48A protein had only a three-fold reduction in endonuclease activity. This research has shown that MutL stimulates the wild-type MutH activity by 1000-fold. The wild-type MutH stimulation by MutL for binding was only shown to be 16-fold. The G49A MutH mutant interferes with the proper functioning of the protein but is not informative about the mechanism of action. The binding ability of this mutant was the same as wild-type and the endonuclease activity was down 30-fold with a 10-fold stimulation by MutL. The extra methyl group of the alanine may cause slight structural changes in the lysine 48 side chain that slows catalysis. The two histidines (H112 and H115) in MutH that are in a similar position as the two histidines (H84 and H85) in PvuII (that signal for DNA binding and catalysis) were changed to alanines, but had wild-type activity both in-vivo and in-vitro. These results indicate that the MutH signal for DNA binding and catalysis remains unknown. The two deletion mutations (MutHΔ224 and MutHΔ214) in the C-terminal end of the protein, localized the MutL stimulation region to five amino acids (Ala220, Leu221, Leu222, Ala223, and Arg224). Mutant MutHΔ224 had wild-type MutL stimulation activity, while MutHΔ214 showed no MutL stimulation. Another deletion mutant, MutHΔ119, from another laboratory was shown to have wild-type MutL stimulation also. This leaves one (or more) of the remaining five residues as important for MutL stimulation.Biochemistry and Molecular Pharmacolog
Urban Structure and Housing Prices: Some Evidence from Australian Cities
No full textThis paper studies determinants of some aspects of the structure of cities, including density and the price of land and housing. We use a version of the Alonso-Muth-Mills model, calibrated to broadly match some of the features of a representative large city. While the calibrated model omits many real-world features, it can nonetheless be used to explore the impact of factors such as: (i) the provision of transport infrastructure; (ii) zoning policies that limit housing density; (iii) frictions on the production of housing; and (iv) population size. The empirical section of the paper shows that the model is consistent with some empirical regularities for large Australian cities. The results of the paper draw attention to structural factors that may have contributed to developments in the Australian housing market in recent years.housing; housing prices; land prices; zoning; land use
DNA mismatch correction in Haemophilus influenzae: characterization of MutL, MutH and their interaction
No full textHaemophilus influenzae DNA mismatch repair proteins, MutS, MutL and MutH, are functionally characterized in this study. Introduction of mutS, mutL and mutH genes of H. influenzae resulted in complementation of the mismatch repair activity of the respective mutant strains of Escherichia coli to varying levels. DNA binding studies using H. influenzae MutH have shown that the protein is capable of binding to any DNA sequence non-specifically in a co-operative and metal independent manner. Presence of MutL and ATP in the binding reaction resulted in the formation of a more specific complex, which indicates that MutH is conferred specificity for binding hemi-methylated DNA through structural alterations mediated by its interaction with MutL. To study the role of conserved amino acids Ile213 and Leu214 in the helix at the C-terminus of MutH, they were mutated to alanine. The mutant proteins showed considerably reduced DNA binding and nicking, as well as MutL-mediated activation. MutH failed to nick HU bound DNA whereas MboI and Sau3AI, which have the same recognition sequence as MutH, efficiently cleaved the substrate. MutS ATPase activity was found to be reduced two-fold in presence of covalently closed circular duplex containing a mismatched base pair whereas, the activity was regained upon linearization of the circular duplex. This observation possibly suggests that the MutS clamps are trapped in the closed DNA heteroduplex. These studies, therefore, serve as the basis for a detailed investigation of the structure-function relationship among the protein partners of the mismatch repair pathway of H. influenzae
DNA mismatch correction in Haemophilus influenzae: characterization of MutL, MutH and their interaction
No full textHaemophilus influenzae DNA mismatch repair proteins, MutS, MutL and MutH, are functionally characterized in this study. Introduction of mutS, mutL and mutH genes of H. influenzae resulted in complementation of the mismatch repair activity of the respective mutant strains of Escherichia coli to varying levels. DNA binding studies using H. influenzae MutH have shown that the protein is capable of binding to any DNA sequence non-specifically in a co-operative and metal independent manner. Presence of MutL and ATP in the binding reaction resulted in the formation of a more specific complex, which indicates that MutH is conferred specificity for binding hemi-methylated DNA through structural alterations mediated by its interaction with MutL. To study the role of conserved amino acids Ile213 and Leu214 in the helix at the C-terminus of MutH, they were mutated to alanine. The mutant proteins showed considerably reduced DNA binding and nicking, as well as MutL-mediated activation. MutH failed to nick HU bound DNA whereas MboI and Sau3AI, which have the same recognition sequence as MutH, efficiently cleaved the substrate. MutS ATPase activity was found to be reduced two-fold in presence of covalently closed circular duplex containing a mismatched base pair whereas, the activity was regained upon linearization of the circular duplex. This observation possibly suggests that the MutS clamps are trapped in the closed DNA heteroduplex. These studies, therefore, serve as the basis for a detailed investigation of the structure-function relationship among the protein partners of the mismatch repair pathway of H. influenzae
Das treue Mädchen weinte Blut : Und dennoch wandelte voll Muth der Heilige von dannen
No full textAbschied des Eynthio von RosetteBoemasler del. ; C. Schule sc. 1808Auf Unterlagenblatt handschriftliche Notiz "nicht Lips"Oben links bezeichnet "I." und unten Mitte "Das treue Mädchen weinte Blut ; / Und dennoch wandelte voll Muth / Der Heilige von dannen." / II.r Thl. S. 203.
The Muth Formation in the Pin Valley (Spiti, N-India)
No full textIm Pin Tal (Spiti) wurden jungproterozoischen und altpaläozoischen Sedimentserien bearbeitet. Lithologische Profile von der Pin Fm., Muth Fm. und Lipak Fm. wurden angefertigt und leicht erkennbare Grenzen zwischen den Formationen definiert. Charakteristische lithologische Variationen ermöglichen eine überregionale Korrelation dieser Serien mit den Tethys Sedimenten von Zanskar bis Kumaon. Die Muth Fm. wird anhand ihrer sedimentären Strukturen als ein Barriereninsel System interpretiert. Basierend auf lithologischen Veränderungen und unterschiedlichen sedimentärer Strukturen, werden 4 Fazies Assoziationen (FA1-FA4) unterschieden. Vom Liegenden ins Hangende finden sich beach, shoreface bis foreshore, Küstendünen, Lagune und abschließend wieder shoreface Ablagerungen. Die Spurenfossilien Vergesellschaftung besteht aus zahlreichen Palmichnium antarcticum und Diplichnites gouldi. Deutlich seltener finden sich Diplopodichnus biformis, Taenidium barretti, Didymaulichnus cf. lyelli, Didymaulyponomos cf. rowei, Metaichnia isp. und verticale Bohrgänge mit unklarer systematischer Zuordnung. Da in der Muth Fm. gut datierbaren Fossilien gefunden wurden, läßt sich lediglich das Mindestalter der Formation anhand der Datierung der überlagernden Lipak Fm. feststellen. Es wurden deshalb aus den ersten Kalkbänken der Lipak Fm. Gut erhaltene Conodonten, etwa 30 m oberhalb der Lipak Basis, zeigen ein mitteldevonisches Alter (Givet) an.
SE von Mikkim wurden zahlreiche deformation bands im gesamten Aufschlußbereich gefunden. Die Rekonstruktion dieser Strukturen auf ihre Orientierung vor der Verfaltung durch die Himalaya Orogenese, zeigt Ost-West streichende, subvertical einfallende Störungen, die eine Deformation anzeigen, die älter ist als die Himalaya Orogenese. Da es sich dabei um Strukturen handelt, die aus porösen Sandsteinen beschrieben sind, liegt das Alter dieser Deformation zwischen dem Sedimentationsalter der Muth Fm. und deren vollständiger Zementation.Uppermost Proterozoic to Lower Paleozoic sedimentary sequences in the Pin Valley (Spiti) have been investigated. Lithological sections of the Pin Fm., Muth Fm. and Lipak Fm. have been measured, and useful, clear recognizable boundaries between the Formations have been defined. Typical lithological variations permit the regional correlation with Tethyan sediments from Zanskar to Kumaon. Sedimentary structures indicate that the Muth Fm. was deposited in a barrier-island system. Based on lithological variations and different sedimentary structures, four facies (FA1-FA4) have been distinguished, comprising (beginning at the base) shoreface to foreshore, coastal dune, lagoon and again shoreface depositional environments. The ichnoassemblage consists of abundant Palmichnium antarcticum and Diplichnites gouldi with rarer Diplopodichnus biformis, Taenidium barretti, Didymaulichnus cf. lyelli, Didymaulyponomos cf. rowei, Metaichnia isp. and vertical burrows of unclear affinity. Due to the lack of age-indicative fossils in the Muth Fm. the lower age limit of the Formation is constrained by the age of the overlying Lipak Fm. Conondont samples have been taken at the lowermost occurrence of limestone beds in the Lipak Fm. (c. 30 m above the base of the Formation), that yielded nicely preserved conodonts of Givetian age. Deformation bands are very common in the Muth Fm. SE of Mikkim. Restored to their original orientation, they represent near vertical, E-W striking faults, indicating pre-Himalayan deformation with an older age limit represented by the age of sedimentation and a younger age limit represented by the age of complete cementation of the arenite
The plasmid DNAs cleavage by MutH in a MutS-MutL dependent manner.
No full text<p><b>A</b>, Analysis of the G/T-cccDNA treated with MutS-MutL-MutH mixture after 1, 5 or 10 min incubation in 1% agarose gel containing ethidium bromide. The initial cccDNA is shown (0 min). M – DNA ladder. <b>B</b>, Diagram representing the data of hydrolysis by MutS-MutL-MutH mixture of G/T-, G/C-, G/Tg- and A/Tg-cccDNA (the variable nucleotide pair introduced in cccDNA is indicated under the lanes) for 5 min. The experiments were performed 5 times. Error bars are standard deviations of the mean.</p
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