scholarly journals Deacylation of acylamino compounds other than penicillins by the cell-bound penicillin acylase of Escherichia coli

1969 ◽  
Vol 115 (4) ◽  
pp. 741-745 ◽  
Author(s):  
M. Cole
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Small Group ◽  
Penicillin Acylase ◽  
E Coli ◽  
Rapid Hydrolysis ◽  
Hydrolysis Of

1. The action of the penicillin acylase enzyme of Escherichia coli N.C.I.B. 8743 on non-penicillin substrates suggests that the enzyme is an amidohydrolase. 2. The rates of hydrolysis for a small group of penicillins closely parallel those for a corresponding series of N-acylglycines. 3. For a series of E. coli strains, ability to cause rapid hydrolysis of phenylacetylglycine is correlated with ability to hydrolyse benzylpenicillin. 4. Amides and N-acylglycines are hydrolysed to the corresponding acids. The phenylacetyl group is hydrolysed most readily. Benzamide and β-phenylpropionamide are not substrates. In a series of aliphatic acylglycines only valeryl- and hexanoyl-glycine are substrates. 5. Acylated l- but not d-α-amino acids are hydrolysed. d-α-Hydroxyphenylacetamide is a better substrate than the l compound.

scholarly journals Penicillins and other acylamino compounds synthesized by the cell-bound penicillin acylase of Escherichia coli

1969 ◽  
Vol 115 (4) ◽  
pp. 747-756 ◽  
Author(s):  
M. Cole
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Phenylacetic Acid ◽  
Enzyme Reaction ◽  
Reaction Rates ◽  
Hydroxamic Acids ◽  
Penicillin Acylase ◽  
E Coli ◽  
Hydroxyphenylacetic Acid ◽  
Derivatives Of

1. The penicillin acylase of Eschericha coli N.C.I.B. 8743 is a reversible enzyme. Reaction rates for the two directions have been determined. 2. Measurements of the rates of enzymic synthesis of penicillins from 6-aminopenicillanic acid and various carboxylic acids revealed that p-hydroxyphenylacetic acid was the best substrate, followed by phenylacetic, 2-thienylacetic, substituted phenylacetic, 3-hexenoic and n-hexanoic acids. 3. The rate of synthesis of penicillin improved when amides or N-acylglycines were used; α-aminobenzylpenicillin and phenoxymethylpenicillin were only synthesized when using these more energy-rich compounds. 4. Phenyl-acetylglycine was the best substrate for the synthesis of benzylpenicillin compared with other derivatives of phenylacetic acid. 5. The enzyme was specific for acyl-l-amino acids, benzylpenicillin being synthesized from phenylacetyl-l-α-aminophenylacetic acid but not from phenylacetyl-d-α-aminophenylacetic acid. 6. α-Phenoxyethylpenicillin was synthesized from 6-aminopenicillanic acid and α-phenoxypropionylthioglycollic acid non-enzymically, but the rate was faster in the presence of the enzyme. 7. The E. coli acylase catalysed the acylation of hydroxylamine by acids or amides to give hydroxamic acids, the phenylacetyl group being the most suitable acyl group. The enzyme also catalysed other acyl-group transfers.

scholarly journals Roles of the C-Terminal Amino Acids of Non-Hexameric Helicases: Insights from Escherichia coli UvrD

2021 ◽  
Vol 22 (3) ◽  
pp. 1018
Author(s):  
Hiroaki Yokota
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Single Molecule ◽  
Underlying Mechanism ◽  
Amino Acid Sequences ◽  
E Coli ◽  
X Ray Crystallography ◽  
Terminal Amino

Helicases are nucleic acid-unwinding enzymes that are involved in the maintenance of genome integrity. Several parts of the amino acid sequences of helicases are very similar, and these quite well-conserved amino acid sequences are termed “helicase motifs”. Previous studies by X-ray crystallography and single-molecule measurements have suggested a common underlying mechanism for their function. These studies indicate the role of the helicase motifs in unwinding nucleic acids. In contrast, the sequence and length of the C-terminal amino acids of helicases are highly variable. In this paper, I review past and recent studies that proposed helicase mechanisms and studies that investigated the roles of the C-terminal amino acids on helicase and dimerization activities, primarily on the non-hexermeric Escherichia coli (E. coli) UvrD helicase. Then, I center on my recent study of single-molecule direct visualization of a UvrD mutant lacking the C-terminal 40 amino acids (UvrDΔ40C) used in studies proposing the monomer helicase model. The study demonstrated that multiple UvrDΔ40C molecules jointly participated in DNA unwinding, presumably by forming an oligomer. Thus, the single-molecule observation addressed how the C-terminal amino acids affect the number of helicases bound to DNA, oligomerization, and unwinding activity, which can be applied to other helicases.

scholarly journals 1D TiO2 Nanostructures Prepared from Seeds Presenting Tailored TiO2 Crystalline Phases and Their Photocatalytic Activity for Escherichia coli in Water

2018 ◽  
Vol 2018 ◽  
pp. 1-6 ◽  
Author(s):  
Julieta Cabrera ◽  
Dwight Acosta ◽  
Alcides López ◽  
Roberto J. Candal ◽  
Claudia Marchi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Photocatalytic Activity ◽  
Bactericidal Activity ◽  
Sol Gel ◽  
Titanium Isopropoxide ◽  
E Coli ◽  
1D Nanostructures ◽  
Sol Gel Synthesis ◽  
Hydrolysis Of ◽  
Tio2 Nanostructures

TiO2 nanotubes were synthesized by alkaline hydrothermal treatment of TiO2 nanoparticles with a controlled proportion of anatase and rutile. Tailoring of TiO2 phases was achieved by adjusting the pH and type of acid used in the hydrolysis of titanium isopropoxide (first step in the sol-gel synthesis). The anatase proportion in the precursor nanoparticles was in the 3–100% range. Tube-like nanostructures were obtained with an anatase percentage of 18 or higher while flake-like shapes were obtained when rutile was dominant in the seed. After annealing at 400°C for 2 h, a fraction of nanotubes was conserved in all the samples but, depending on the anatase/rutile ratio in the starting material, spherical and rod-shaped structures were also observed. The photocatalytic activity of 1D nanostructures was evaluated by measuring the deactivation of E. coli in stirred water in the dark and under UV-A/B irradiation. Results show that in addition to the bactericidal activity of TiO2 under UV-A illumination, under dark conditions, the decrease in bacteria viability is ascribed to mechanical stress due to stirring.

Increased Production of Recombinant O-Phospho-L-Serine Sulfhydrylase from the Hyperthermophilic Archaeon Aeropyrum pernix K1 Using Escherichia coli

2019 ◽  
Vol 8 (1) ◽  
pp. 15-23
Author(s):  
Takashi Nakamura ◽  
Emi Takeda ◽  
Tomoko Kiryu ◽  
Kentaro Mori ◽  
Miyu Ohori ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Industrial Production ◽  
Codon Optimization ◽  
Scale Production ◽  
Enzymatic Production ◽  
Hyperthermophilic Archaeon ◽  
E Coli ◽  
Aeropyrum Pernix ◽  
Natural Amino Acids

Background: O-phospho-L-serine sulfhydrylase from the hyperthermophilic archaeon Aeropyrum pernix K1 (ApOPSS) is thermostable and tolerant to organic solvents. It can produce nonnatural amino acids in addition to L-cysteine. Objective: We aimed to obtain higher amounts of ApOPSS compared to those reported with previous methods for the convenience of research and for industrial production of L-cysteine and non-natural amino acids. Method: We performed codon optimization of cysO that encodes ApOPSS, for optimal expression in Escherichia coli. We then examined combinations of conditions such as the host strain, plasmid, culture medium, and isopropyl β-D-1-thiogalactopyranoside (IPTG) concentration to improve ApOPSS yield. Results and Discussion: E. coli strain Rosetta (DE3) harboring the expression plasmid pQE-80L with the codon-optimized cysO was cultured in Terrific broth with 0.01 mM IPTG at 37°C for 48 h to yield a 10-times higher amount of purified ApOPSS (650 mg·L-1) compared to that obtained by the conventional method (64 mg·L-1). We found that the optimal culture conditions along with codon optimization were essential for the increased ApOPSS production. The expressed ApOPSS had a 6-histidine tag at the N-terminal, which did not affect its activity. This method may facilitate the industrial production of cysteine and non-natural amino acids using ApOPSS. Conclusion: We conclude that these results could be used in applied research on enzymatic production of L-cysteine in E. coli, large scale production of non-natural amino acids, an enzymatic reaction in organic solvent, and environmental remediation by sulfur removal.

scholarly journals Site-Specific Incorporation of Unnatural Amino Acids into Escherichia coli Recombinant Protein: Methodology Development and Recent Achievement

Biomolecules ◽  
2019 ◽  
Vol 9 (7) ◽  
pp. 255 ◽  
Author(s):  
Sviatlana Smolskaya ◽  
Yaroslav Andreev
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Unnatural Amino Acids ◽  
Trna Synthetase ◽  
Nonsense Suppression ◽  
Expression Vectors ◽  
Site Specific ◽  
Suppressor Trna ◽  
E Coli ◽  
Orthogonal Pair

More than two decades ago a general method to genetically encode noncanonical or unnatural amino acids (NAAs) with diverse physical, chemical, or biological properties in bacteria, yeast, animals and mammalian cells was developed. More than 200 NAAs have been incorporated into recombinant proteins by means of non-endogenous aminoacyl-tRNA synthetase (aa-RS)/tRNA pair, an orthogonal pair, that directs site-specific incorporation of NAA encoded by a unique codon. The most established method to genetically encode NAAs in Escherichia coli is based on the usage of the desired mutant of Methanocaldococcus janaschii tyrosyl-tRNA synthetase (MjTyrRS) and cognate suppressor tRNA. The amber codon, the least-used stop codon in E. coli, assigns NAA. Until very recently the genetic code expansion technology suffered from a low yield of targeted proteins due to both incompatibilities of orthogonal pair with host cell translational machinery and the competition of suppressor tRNA with release factor (RF) for binding to nonsense codons. Here we describe the latest progress made to enhance nonsense suppression in E. coli with the emphasis on the improved expression vectors encoding for an orthogonal aa-RA/tRNA pair, enhancement of aa-RS and suppressor tRNA efficiency, the evolution of orthogonal EF-Tu and attempts to reduce the effect of RF1.

Inhibition of Escherichia coli O157:H7 in Ripening Dry Fermented Sausage by Ground Yellow Mustard

2008 ◽  
Vol 71 (3) ◽  
pp. 486-493 ◽  
Author(s):  
GARY H. GRAUMANN ◽  
RICHARD A. HOLLEY
Keyword(s):  
Escherichia Coli ◽  
Starter Culture ◽  
Escherichia Coli O157 ◽  
Fermented Sausage ◽  
Yellow Mustard ◽  
E Coli ◽  
Dry Fermented Sausage ◽  
Hydrolysis Of ◽  
Antimicrobial Effectiveness ◽  
Dry Sausage

Compounds generated by the enzymatic hydrolysis of glucosinolates naturally present in mustard powder are potently bactericidal against Escherichia coli O157:H7. Because E. coli O157:H7 can survive the dry fermented sausage manufacturing process, 2, 4, and 6% (wt/wt) nondeheated (hot) mustard powder or 6% (wt/wt) deheated (cold) mustard powder were added to dry sausage batter inoculated with E. coli O157:H7 at about 7 log CFU/g to evaluate the antimicrobial effectiveness of the powders. Reductions in E. coli O157:H7 populations, changes in pH and water activity (aw), effects on starter culture (Pediococcus pentosaceus and Staphylococcus carnosus) populations, and effects of mustard powder on sausage texture (shear) were monitored during ripening. Nondeheated mustard powder at 2, 4, and 6% in dry sausage (0.90 aw) resulted in significant reductions in E. coli O157:H7 (P < 0.05) of 3.4, 4.4, and 6.9 log CFU/g, respectively, within 30 days of drying. During fermentation and drying, mustard powder did not affect P. pentosaceus and S. carnosus activity in any of the treatments. Extension of drying to 36 and 48 days reduced E. coli O157:H7 by >5 log CFU/g in the 4 and 2% mustard powder treatments, respectively. The 6% deheated mustard powder treatment provided the most rapid reductions of E. coli O157:H7 (yielding <0.20 log CFU/g after 24 days) by an unknown mechanism and was the least detrimental (P < 0.05) to sausage texture.

Iron(III) complexes of DL-methionine

1972 ◽  
Vol 25 (2) ◽  
pp. 421 ◽  
Author(s):  
EJ Halbert ◽  
MJ Rogerson
Keyword(s):  
Aqueous Solution ◽  
Amino Acids ◽  
Large Excess ◽  
Ph Titration ◽  
Oxidation Reduction ◽  
Infrared Studies ◽  
Rapid Hydrolysis ◽  
Iron Analysis ◽  
Hydrolysis Of ◽  
Made In

Few iron(111) complexes of amino acids have been isolated although attempts have been made to measure their stabilities in solution. Iron(111) in the presence of various amino acids showed no complex formation during pH titration in aqueous solution. Using oxidation-reduction and spectrophotometric techniques Perrin measured the stabilities of 1 : 1 complexes of iron(111) with different amino acids in solutions of low pH. Rapid hydrolysis of iron(111) occurred when the pH was increased even in the presence of a large excess of amino acids. Bielig and Bayer reported the isolation of a bis-complex of iron(111) and methionine prepared in aqueous solution. McAuliffe, Quagliano, and Vallarinoe reported a tris-complex made in ethanol, though their iron analysis was not consistent with the required structure. In both cases magnetic and infrared studies were used to characterize the products. Our attempts to repeat these preparations were unsuccessful. This paper describes the preparation of two 1 : 1 complexes of methionine and iron(111) and an investigation of their properties by analysis of infrared, magnetic, and N.M.R. measurements.

Changes in the permeability of Escherichia coli during parasitization by Bdellovibrio bacteriovorus

1971 ◽  
Vol 17 (5) ◽  
pp. 689-697 ◽  
Author(s):  
S. F. Crothers ◽  
J. Robinson
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Host Cell ◽  
Culture Fluid ◽  
Infection Cycle ◽  
Bdellovibrio Bacteriovorus ◽  
E Coli ◽  
Hepes Buffer ◽  
Per Se ◽  
Ph Range

Bdellovibrio bacteriovorus strain 6-5-S completed a typical infection cycle when incubated with E. coli ML 35 (lac i−z+y−) in 0.025 M Hepes buffer, pH 7.8, supplemented with 0.002 M CaCl2∙2H2O. Growth of this strain of B. bacteriovorus was optimal over the range of pH 7.5–8.5. No growth occurred at pH 6.5. The broad pH range may occur because a buffer per se is not required for growth and multiplication. The parasite failed to multiply in a two-membered culture unsupplemented with Ca2+ or Mg2+.Growth and multiplication of B. bacteriovorus in a two-membered culture were assessed by various parameters, including decrease in absorbance at 520 mμ, and release of materials absorbing at 260 mμ, and at 280 mμ. The onset of lysis of the host cell was accompanied by an increase in the release of materials absorbing at 260 mμ and at 280 mμ. The ratio of the absorbance of these materials at 280 and 260 mμ increased at the same time, from which it may be inferred that probably amino acids or proteins were being released.No β-galactosidase could be detected in the culture fluid of the two-membered culture. The infected E. coli cells were more permeable than uninfected cells to o-nitrophenyl-β-D-galactoside, and to the fluorescent dye 8-aniIino-1-naphthalenesulfonic acid.

scholarly journals Glutathione-Dependent Conversion ofN-Ethylmaleimide to the Maleamic Acid by Escherichia coli: an Intracellular Detoxification Process

2000 ◽  
Vol 66 (4) ◽  
pp. 1393-1399 ◽  
Author(s):  
D. McLaggan ◽  
H. Rufino ◽  
M. Jaspars ◽  
I. R. Booth
Keyword(s):  
Escherichia Coli ◽  
Time Course ◽  
E Coli ◽  
Transient Activation ◽  
Low Concentrations ◽  
Maleamic Acid ◽  
High Concentrations ◽  
Hydrolysis Of ◽  
Strong Activator ◽  
Detoxification Process

ABSTRACT The electrophile N-ethylmaleimide (NEM) elicits rapid K+ efflux from Escherichia coli cells consequent upon reaction with cytoplasmic glutathione to form an adduct, N-ethylsuccinimido-S-glutathione (ESG) that is a strong activator of the KefB and KefC glutathione-gated K+ efflux systems. The fate of the ESG has not previously been investigated. In this report we demonstrate that NEM andN-phenylmaleimide (NPM) are rapidly detoxified by E. coli. The detoxification occurs through the formation of the glutathione adduct of NEM or NPM, followed by the hydrolysis of the imide bond after which N-substituted maleamic acids are released. N-Ethylmaleamic acid is not toxic to E. coli cells even at high concentrations. The glutathione adducts are not released from cells, and this allows glutathione to be recycled in the cytoplasm. The detoxification is independent of new protein synthesis and NAD+-dependent dehydrogenase activity and entirely dependent upon glutathione. The time course of the detoxification of low concentrations of NEM parallels the transient activation of the KefB and KefC glutathione-gated K+ efflux systems.

scholarly journals Enzymic synthesis of N-acetyl-l-phenylalanine in Escherichia coli K12

1971 ◽  
Vol 124 (5) ◽  
pp. 905-913 ◽  
Author(s):  
R. V. Krishna ◽  
P. R. Krishnaswamy ◽  
D. Rajagopal Rao
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Substrate Specificity ◽  
Acetyl Coa ◽  
Ph Optimum ◽  
E Coli ◽  
Enzymic Synthesis ◽  
Escherichia Coli K12

1. Cell-free extracts of Escherichia coli K12 catalyse the synthesis of N-acetyl-l-phenylalanine from acetyl-CoA and l-phenylalanine. 2. The acetyl-CoA–l-phenylalanine α-N-acetyltransferase was purified 160-fold from cell-free extracts. 3. The enzyme has a pH optimum of 8 and catalyses the acetylation of l-phenylalanine. Other l-amino acids such as histidine and alanine are acetylated at slower rates. 4. A transacylase was also purified from E. coli extracts and its substrate specificity studied. 5. The properties of both these enzymes were compared with those of other known amino acid acetyltransferases and transacylases.

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