scholarly journals Hydrolysis of penicillins and related compounds by the cell-bound penicillin acylase of Escherichia coli

1969 ◽  
Vol 115 (4) ◽  
pp. 733-739 ◽  
Author(s):  
M. Cole
Keyword(s):  
Escherichia Coli ◽  
Penicillin Acylase ◽  
Side Chain ◽  
Bacterial Cells ◽  
Optimum Temperature ◽  
Phenoxymethyl Penicillin ◽  
Optimum Ph ◽  
Penicillanic Acid ◽  
Hydrolysis Of ◽  
Related Compounds

1. A method is given for the preparation of penicillin acylase by using Escherichia coli N.C.I.B. 8743 and a strain selected for higher yield. The enzyme is associated with the bacterial cells and removes the side chains of penicillins to give 6-amino-penicillanic acid and a carboxylic acid. 2. The rates of penicillin deacylation indicated that p-hydroxybenzylpenicillin was the best substrate, followed in diminishing order by benzyl-, dl-α-hydroxybenzyl-, 2-furylmethyl-, 2-thienylmethyl-, d-α-aminobenzyl-, n-propoxymethyl- and isobutoxymethyl-penicillin. Phenylpenicillin and dl-α-carboxybenzylpenicillin were not substrates and phenoxymethyl-penicillin was very poor. 3. Amides and esters of the above penicillins were also substrates for the deacylation reaction, as were cephalosporins with a thienylmethyl side chain. 4. For the deacylation of 2-furylmethylpenicillin at 21° the optimum pH was 8·2. The optimum temperature was 60° at pH7. 5. By using selection A of N.C.I.B. 8743 and determining reaction velocities by assaying yields of 6-amino-penicillanic acid in a 10min. reaction at 50° and pH8·2, the Km for benzylpenicillin was found to be about 30mm and the Km for 2-furylmethylpenicillin, about 10mm. The Vmax. values were 0·6 and 0·24μmole/min./mg. of bacterial cells respectively.

scholarly journals Properties of a new fungal b-galactosidase with potential application in the dairy industry

1999 ◽  
Vol 30 (3) ◽  
pp. 265-271 ◽  
Author(s):  
Rubens Cruz ◽  
Vinícius D'Arcádia Cruz ◽  
Juliana Gisele Belote ◽  
Marcelo de Oliveira Khenayfes ◽  
Claudia Dorta ◽  
...  
Keyword(s):  
Hydrolytic Activity ◽  
Industrial Applications ◽  
Transferase Activity ◽  
Optimum Temperature ◽  
Milk Whey ◽  
Beneficial Effects ◽  
Optimum Ph ◽  
Semi Solid ◽  
Good Productivity ◽  
Hydrolysis Of

<FONT FACE="Symbol">b</font>-Galactosidase or <FONT FACE="Symbol">b</font>-D-galactoside-galactohydrolase (EC. 3.2.1.23) is an important enzyme industrially used for the hydrolysis of lactose from milk and milk whey for several applications. Lately, the importance of this enzyme was enhanced by its galactosyltransferase activity, which is responsible for the synthesis of transgalactosylated oligosaccharides (TOS) that act as functional foods, with several beneficial effects on consumers. Penicillium simplicissimum, a strain isolated from soil, when grown in semi-solid medium showed good productivity of <FONT FACE="Symbol">b</font>-galactosidase with galactosyltransferase activity. The optimum pH for hydrolysis was in the 4.04.6 range and the optimum pH for galactosyltransferase activity was in the 6.07.0 range. The optimum temperature for hydrolysis and transferase activity was 55-60°C and 50°C, respectively, and the enzyme showed high thermostability for the hydrolytic activity. The enzyme showed a potential for several industrial applications such as removal of 67% of the lactose from milk and 84% of the lactose from milk whey when incubated at their original pH (4.5 and 6.34, respectively) under optimum temperature conditions. When incubated with a 40% lactose solution in 150 mM McIlvaine buffer, pH 4.5, at 55°C the enzyme converted 86.5% of the lactose to its component monosaccharides. When incubated with a 60% lactose solution in the same buffer but at pH 6.5 and 50°C, the enzyme can synthetize up to 30.5% TOS, with 39.5% lactose and 30% monosaccharides remaining in the preparation.

scholarly journals Purification and Substrate Specificities of Two α-l-Arabinofuranosidases from Aspergillus awamori IFO 4033

1998 ◽  
Vol 64 (10) ◽  
pp. 4021-4027 ◽  
Author(s):  
Satoshi Kaneko ◽  
Mitsue Arimoto ◽  
Misako Ohba ◽  
Hideyuki Kobayashi ◽  
Tadashi Ishii ◽  
...  
Keyword(s):  
Culture Filtrate ◽  
The Other ◽  
Hydrolysis Reaction ◽  
Aspergillus Awamori ◽  
Side Chain ◽  
Optimum Temperature ◽  
Substrate Specificities ◽  
Molecular Weights ◽  
Ph Values ◽  
Optimum Ph

ABSTRACT α-l-Arabinofuranosidases I and II were purified from the culture filtrate of Aspergillus awamori IFO 4033 and had molecular weights of 81,000 and 62,000 and pIs of 3.3 and 3.6, respectively. Both enzymes had an optimum pH of 4.0 and an optimum temperature of 60°C and exhibited stability at pH values from 3 to 7 and at temperatures up to 60°C. The enzymes released arabinose from p-nitrophenyl-α-l-arabinofuranoside,O-α-l-arabinofuranosyl-(1→3)-O-β-d-xylopyranosyl-(1→4)-d-xylopyranose, and arabinose-containing polysaccharides but not fromO-β-d-xylopyranosyl-(1→2)-O-α-l-arabinofuranosyl-(1→3)-O-β-d-xylopyranosyl-(1→4)-O-β-d-xylopyranosyl-(1→4)-d-xylopyranose. α-l-Arabinofuranosidase I also released arabinose fromO-β-d-xylopy-ranosyl-(1→4)-[O-α-l-arabinofuranosyl-(1→3)]-O-β-d-xylopyranosyl-(1→4)-d-xylopyranose. However, α-l-arabinofuranosidase II did not readily catalyze this hydrolysis reaction. α-l-Arabinofuranosidase I hydrolyzed all linkages that can occur between two α-l-arabinofuranosyl residues in the following order: (1→5) linkage > (1→3) linkage > (1→2) linkage. α-l-Arabinofuranosidase II hydrolyzed the linkages in the following order: (1→5) linkage > (1→2) linkage > (1→3) linkage. α-l-Arabinofuranosidase I preferentially hydrolyzed the (1→5) linkage of branched arabinotrisaccharide. On the other hand, α-l-arabinofuranosidase II preferentially hydrolyzed the (1→3) linkage in the same substrate. α-l-Arabinofuranosidase I released arabinose from the nonreducing terminus of arabinan, whereas α-l-arabinofuranosidase II preferentially hydrolyzed the arabinosyl side chain linkage of arabinan.

scholarly journals Biosynthesis of Putrescine from L-arginine Using Engineered Escherichia coli Whole Cells

Catalysts ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 947
Author(s):  
Hongjie Hui ◽  
Yajun Bai ◽  
Tai-Ping Fan ◽  
Xiaohui Zheng ◽  
Yujie Cai
Keyword(s):  
Escherichia Coli ◽  
Biogenic Amine ◽  
Arginine Decarboxylase ◽  
Optimal Concentration ◽  
Optimum Temperature ◽  
Whole Cell ◽  
Whole Cells ◽  
Copy Numbers ◽  
Optimum Ph ◽  
Engineered Escherichia Coli

Putrescine, a biogenic amine, is a highly valued compound in medicine, industry, and agriculture. In this study, we report a whole-cell biocatalytic method in Escherichia coli for the production of putrescine, using L-arginine as the substrate. L-arginine decarboxylase and agmatine ureohydrolase were co-expressed to produce putrescine from L-arginine. Ten plasmids with different copy numbers and ordering of genes were constructed to balance the expression of the two enzymes, and the best strain was pACYCDuet-speB-speA. The optimal concentration of L-arginine was determined to be 20 mM for this strain. The optimum pH of the biotransformation was 9.5, and the optimum temperature was 45 °C; under these conditions, the yield of putrescine was 98%. This whole-cell biocatalytic method appeared to have great potential for the production of putrescine.

Multiresidue Method for Determination of Eight Neutral β-Lactam Penicillins in Milk by Fluorescence-Liquid Chromatography

1985 ◽  
Vol 68 (5) ◽  
pp. 968-971
Author(s):  
Robert K Munns ◽  
Wilbert Shimoda ◽  
Jose E Roybal ◽  
Cassandra Vieira
Keyword(s):  
Liquid Chromatography ◽  
Mobile Phase ◽  
Methylene Chloride ◽  
Side Chain ◽  
Excitation Wavelength ◽  
Benzyl Penicillin ◽  
Aqueous Fraction ◽  
Phenoxymethyl Penicillin ◽  
Hydrolysis Of

Abstract A method of determining total penicillins begins with an enzymatic hydrolysis of the (J-Iactam ring to form their respective penicilloate product. Acetonitrile precipitates much of the casein and protein, which are then separated from the liquid by centrifugation. The lipids are removed from the aqueous fraction with methylene chloride. Mercuric chloride is added, which reacts with the penicilloate to liberate the side chain that has a terminal aldehyde. These penilloaldehyde products are extracted with methylene chloride and are subsequently reacted with dansyl hydrazine. The resulting fluorolabeled side chains are separated by liquid chromatography on a C18 column with acetonitrile- water as mobile phase. The fluorescence is measured by the mercury line at 254 nm excitation wavelength and a 500 nm filter on the emission side. The overall average recoveries from milk spiked at 25, 50, and 100 ppb are benzyl penicillin 79.4%; phenoxymethyl penicillin 59.7%; phenethicillin 75.9%; nafcillin 87.7%; methacillin 47.5%; oxacillin 57.6%; cloxicillin 37.3%; and dicloxicillin 26.4%.

scholarly journals Promising Pathway of Thermostable Mannitol Dehydrogenase (MtDH) from Caldicellulosiruptor hydrothermalis 108 for D-Mannitol Synthesis

Separations ◽  
2021 ◽  
Vol 8 (6) ◽  
pp. 76
Author(s):  
Marwa Y. F. Koko ◽  
Rokayya Sami ◽  
Bertrand Muhoza ◽  
Ebtihal Khojah ◽  
Ahmed M. A. Mansour
Keyword(s):  
Escherichia Coli ◽  
Formate Dehydrogenase ◽  
Enzyme System ◽  
Catalytic Efficiency ◽  
Optimum Temperature ◽  
Initial Activity ◽  
Mannitol Dehydrogenase ◽  
Whole Cells ◽  
Optimum Ph ◽  
Molecular Masses

In this study, we conducted the characterization and purification of the thermostable mannitol dehydrogenase (MtDH) from Caldicellulosiruptor hydrothermalis 108. Furthermore, a coupling-enzyme system was designed using (MtDH) from Caldicellulosiruptor hydrothermalis 108 and formate dehydrogenase (FDH) from Ogataea parapolymorpha. The biotransformation system was constructed using Escherichia coli whole cells. The purified enzyme native and subunit molecular masses were 76.7 and 38 kDa, respectively, demonstrating that the enzyme was a dimer. The purified and couple enzyme system results were as follows; the optimum pH for the reduction and the oxidation was 7.0 and 8.0, the optimum temperature was 60 °C, the enzyme activity was inhibited by EDTA and restored by zinc. Additionally, no activity was detected with NADPH and NADP. The purified enzyme showed high catalytic efficiency Kcat 385 s−1, Km 31.8 mM, and kcat/Km 12.1 mM−1 s−1 for D-fructose reduction. Moreover, the purified enzyme retained 80%, 75%, 60%, and 10% of its initial activity after 4 h at 55, 60, 65, and 75 °C, respectively. D-mannitol yield was achieved via HPLC. Escherichia coli are the efficient biotransformation mediator to produce D-mannitol (byproducts free) at high temperature and staple pH, resulting in a significant D-mannitol conversation (41 mg/mL) from 5% D-fructose.

scholarly journals Biochemical Characterization of TEM-92 Extended-Spectrum β-Lactamase, a Protein Differing from TEM-52 in the Signal Peptide

2002 ◽  
Vol 46 (12) ◽  
pp. 3981-3983 ◽  
Author(s):  
Mariagrazia Perilli ◽  
Bernardetta Segatore ◽  
Maria Rosaria De Massis ◽  
Laura Pagani ◽  
Francesco Luzzaro ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Kinetic Parameters ◽  
Signal Peptide ◽  
Proteus Mirabilis ◽  
Biochemical Characterization ◽  
Extended Spectrum ◽  
Penicillanic Acid ◽  
Hydrolysis Of

ABSTRACT A bla TEM-92 gene was cloned from a Proteus mirabilis isolate and expressed in Escherichia coli. Production of the enzyme caused reduction of susceptibility to penicillins and narrow- to expanded-spectrum cephalosporins but not to moxalactam and cephamycins. Determination of kinetic parameters with the purified enzyme revealed hydrolysis of expanded-spectrum cephalosporins, while cephamycins, moxalactam, and aztreonam were very poorly or not hydrolyzed. Clavulanate and penicillanic acid sulfones acylated TEM-92 slowly, and deacylation occurred at measurable rates.

Preparation of Genipin by Hydrolysis of Geniposide with Co-Immobilized Enzyme

2011 ◽  
Vol 236-238 ◽  
pp. 1793-1798 ◽  
Author(s):  
Hua Zheng Liang ◽  
He Chen ◽  
Jian Feng Wang ◽  
Yu Lan He
Keyword(s):  
High Performance ◽  
Immobilized Enzyme ◽  
Low Cost ◽  
Enzymatic Properties ◽  
High Yield ◽  
Cross Linking ◽  
Optimum Temperature ◽  
Environmental Friendly ◽  
Optimum Ph ◽  
Hydrolysis Of

Co-immobilize enzyme by cross-linking and embedding, optimize conditions for immobilizing, determinate the enzymatic properties of co-immobilized enzyme and study the methods for preparation of genipin using co-immobilized enzyme to hydrolyze geniposide. Optimized immobilizing conditions include glutaraldehyde concentration being 0.15%, cross-linking temperature being 20°C, cross-linking time being 2 hours, the activity of co-immobilized β-glucosidase and cell reaches to 65.33U/mg and the enzyme activity recovery being 52.63%. Enzymatic properties of co-immobilized enzyme are following: optimum temperature is 55°C and optimum pH is 5.0. The transformation experiments are carried out with co-immobilized enzyme. The results show that half-life of co-immobilized enzyme reaches around 40 days, higher than the normal immobilized enzyme. The conversion rate of geniposide is above 95% after 8 hours. The genipin is isolated, purified and recrystallized to reach more than 98% of purity by High Performance Liquid Chromatography. Advantages to prepare genipin using co-immobilized enzyme include low cost, high yield, environmental friendly and easy to manufacturing.

scholarly journals Isolasi Mikroorganisme Potensial Penghasil Lipase dari Limbah Pengolahan Minyak Kelapa Sawit Malinping

2020 ◽  
Vol 13 (2) ◽  
pp. 228-241
Author(s):  
Ika Rahmatul Layly ◽  
Erma Widyasti ◽  
Deden Rosid Waltam ◽  
Ayi Mufti ◽  
Nita Wiguna ◽  
...  
Keyword(s):  
Crude Oil ◽  
Fats And Oils ◽  
Optimum Temperature ◽  
Processing Waste ◽  
Titration Method ◽  
Optimum Ph ◽  
Oil Tank ◽  
Textile Industries ◽  
Hydrolysis Of ◽  
Optimum Ph And Temperature

AbstrakLipase adalah kelompok enzim yang mengkatalisis hidrolisis rantai panjang trigliserida, lemak, dan minyak menjadi gliserol dan asam lemak dengan adanya air. Sumber lipase untuk industri kebanyakan berasal dari mikroorganisme. Penggunaan lipase pada industri makin meningkat setiap tahunnya meliputi aplikasinya pada industri makanan, pakan, farmasi, pulp, dan kertas, biodiesel, dan industri tekstil. Dalam usaha mendapatkan isolat potensial penghasil lipase untuk hHidrofilisasi serat poliester, pada penelitian ini dilakukan skrining dan isolasi mikroorganisme yang dapat menghasilkan lipase dari limbah pengolahan minyak kelapa sawit di Malinping, Lebak, Banten. Sebanyak 20 isolat bakteri dan 5 isolat jamur yang diperoleh kemudian diuji aktivitas lipasenya menggunakan metode titrasi. Empat isolat bakteri terpilih (Kondensat, Lumpur-Got, Hasil-Buangan, dan Tangki-Crude-Oil) serta lima isolat jamur (Nut-A, Nut-B, Nut-C, Kernel-B, dan Kernel-C) dikarakterisasi pH dan suhu optimum enzimnya. Hasil karakterisasi pH menunjukkan bahwa isolat bakteri Kondensat, Lumpur-Got, Hasil-Buangan, dan Tangki-Crude-Oil mempunyai aktivitas enzim lipase tertinggi pada pH 6. Suhu optimal aktivitas enzim lipase isolat Lumpur-Got-B, Hasil Buangan-B, dan Tangki-Crude-Oil B  pada 40 °°C, sedangkan isolat bakteri-Kondensat-B optimal pada suhu 30 °°C. Aktivitas lipase kelima isolat jamur optimal pada pH 6. Suhu optimal aktivitas lipase isolat jamur Nut-A adalah 40 °°C, sedangkan isolat Nut-B, Nut-C, Kernel-B, dan Kernel-C aktivitasnya optimal pada 50 °°C.Abstract Lipase are enzymes that catalyzed the hydrolysis of triglyceride, fats and oils into glycerol and fatty acids in the presence of water. Industrial Lipase source mostly derived from microbes. Each year, the lipase utilization in industry increased, such as application for foods, feeds, pharmacys, pulp and papers, biodiesel, and textile industries. On this study, a total of 20 bacteria and 5 fungi lipase potential producer were screened and isolated from oil palm processing waste in Malinping, Lebak, Banten, which then tested for its activity using titration method. Selected isolates then were characterized for its enzyme optimum pH and temperature. The optimum pH for isolate Kondensat, Lumpur-Got, Hasil-Buangan and Crude-Oil-Tank lipases are at pH 6, whilst the optimum temperature of isolates Lumpur-Got B, Hasil-Buangan B and Crude-Oil-Tank B were at 40 °°C and bakteri-Kondensat B isolate optimum at 30 °°C. The five fungi characterization shown optimum pH at 6 and 50 °°C except for isolate Nut-A that optimum at 30 °°C.

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.

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