Hydrolysis of phenylacetanilides catalyzed by penicillin G acylase from Alcaligenes faecalis: Sensitivity of the reaction to substitution in the leaving group

2009 ◽  
Vol 11 (3) ◽  
pp. 196-201 ◽  
Author(s):  
D. Zhiryakova ◽  
M. Guncheva ◽  
I. Ivanov ◽  
N. Stambolieva
Keyword(s):  
Alcaligenes Faecalis ◽  
Penicillin G Acylase ◽  
Penicillin G ◽  
Leaving Group ◽  
Hydrolysis Of

scholarly journals Gold Nanoparticles for Colorimetric detection of hydrolysis of antibiotics by penicillin G acylase

2010 ◽  
Vol 01 (04) ◽  
pp. 322-329 ◽  
Author(s):  
Neha R. Tiwari ◽  
Ambrish Rathore ◽  
Asmita Prabhune ◽  
Sulabha K. Kulkarni
Keyword(s):  
Gold Nanoparticles ◽  
Colorimetric Detection ◽  
Penicillin G Acylase ◽  
Penicillin G ◽  
Hydrolysis Of

Efficient enzymatic synthesis of ampicillin by mutant Alcaligenes faecalis penicillin G acylase

2015 ◽  
Vol 199 ◽  
pp. 62-68 ◽  
Author(s):  
Senwen Deng ◽  
Erzheng Su ◽  
Xiaoqiang Ma ◽  
Shengli Yang ◽  
Dongzhi Wei
Keyword(s):  
Enzymatic Synthesis ◽  
Alcaligenes Faecalis ◽  
Penicillin G Acylase ◽  
Penicillin G

scholarly journals Processing and functional display of the 86 kDa heterodimeric penicillin G acylase on the surface of phage fd

1999 ◽  
Vol 342 (2) ◽  
pp. 415-422 ◽  
Author(s):  
Raymond M. D. VERHAERT ◽  
Jan VAN DUIN ◽  
Wim J. QUAX
Keyword(s):  
Coat Protein ◽  
Signal Sequence ◽  
Protein A ◽  
Alcaligenes Faecalis ◽  
Penicillin Acylase ◽  
Penicillin G Acylase ◽  
Penicillin G ◽  
Β Subunit ◽  
Extracellular Medium ◽  
Α Subunit

The large heterodimeric penicillin G acylase from Alcaligenes faecalis was displayed on the surface of phage fd. We fused the coding sequence (α subunit-internal peptide-β subunit) to the gene of a phage coat protein. A modified g3p signal sequence was used to direct the polypeptide to the periplasm. Here we show that a heterodimeric enzyme can be expressed as a fusion protein that matures to an active biocatalyst connected to the coat protein of phage fd, resulting in a phage to which the β-subunit is covalently linked and the α-subunit is non-covalently attached. The enzyme can be displayed either fused to the minor coat protein g3p or fused to the major coat protein g8p. In both cases the penicillin G acylase on the phage has the same Michaelis constant as its freely soluble counterpart, indicating a proper folding and catalytic activity of the displayed enzyme. The display of the heterodimer on phage not only allows its further use in protein engineering but also offers the possibility of applying this technology for the excretion of the enzyme into the extracellular medium, facilitating purification of the protein. With the example of penicillin acylase the upper limit for a protein to become functionally displayed by phage fd has been further explored. Polyvalent display was not observed despite the use of genetic constructs designed for this aim. These results are discussed in relation to the pore size being formed by the g4p multimer.

Hydrolysis of Penicillin G by Penicillin G Acylase Immobilized on Chitosan Microbeads in Different Reactor Systems

2011 ◽  
Vol 34 (10) ◽  
pp. 1706-1714 ◽  
Author(s):  
M. G. Žuža ◽  
B. M. Obradović ◽  
Z. D. Knežević-Jugović
Keyword(s):  
Penicillin G Acylase ◽  
Penicillin G ◽  
Hydrolysis Of

Asymmetric hydrolysis of dimethyl phenylmalonate by immobilized penicillin G acylase from E. coli

2007 ◽  
Vol 40 (5) ◽  
pp. 997-1000 ◽  
Author(s):  
Zaida Cabrera ◽  
Fernando Lopez-Gallego ◽  
Gloria Fernandez-Lorente ◽  
Jose M. Palomo ◽  
Tamara Montes ◽  
...  
Keyword(s):  
Penicillin G Acylase ◽  
Penicillin G ◽  
E Coli ◽  
Immobilized Penicillin G Acylase ◽  
Hydrolysis Of

scholarly journals Cloning, Overexpression, and Characterization of a Novel Thermostable Penicillin G Acylase from Achromobacter xylosoxidans: Probing the Molecular Basis for Its High Thermostability

2004 ◽  
Vol 70 (5) ◽  
pp. 2764-2770 ◽  
Author(s):  
Gang Cai ◽  
Songcheng Zhu ◽  
Sheng Yang ◽  
Guoping Zhao ◽  
Weihong Jiang
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Molecular Basis ◽  
Alcaligenes Faecalis ◽  
Amino Acid Identity ◽  
Penicillin G Acylase ◽  
Penicillin G ◽  
Achromobacter Xylosoxidans ◽  
High Thermostability ◽  
Arginine Residues

ABSTRACT The gene encoding a novel penicillin G acylase (PGA), designated pgaW, was cloned from Achromobacter xylosoxidans and overexpressed in Escherichia coli. The pgaW gene contains an open reading frame of 2,586 nucleotides. The deduced protein sequence encoded by pgaW has about 50% amino acid identity to several well-characterized PGAs, including those of Providencia rettgeri, Kluyvera cryocrescens, and Escherichia coli. Biochemical studies showed that the optimal temperature for this novel PGA (PGA650) activity is greater than 60°C and its half-life of inactivation at 55°C is four times longer than that of another previously reported thermostable PGA from Alcaligenes faecalis (R. M. D. Verhaert, A. M. Riemens, J. V. R. Laan, J. V. Duin, and W. J. Quax, Appl. Environ. Microbiol. 63:3412-3418, 1997). To our knowledge, this is the most thermostable PGA ever characterized. To explore the molecular basis of the higher thermostability of PGA650, homology structural modeling and amino acid composition analyses were performed. The results suggested that the increased number of buried ion pair networks, lower N and Q contents, excessive arginine residues, and remarkably high content of proline residues in the structure of PGA650 could contribute to its high thermostability. The unique characteristic of higher thermostability of this novel PGA provides some advantages for its potential application in industry.

scholarly journals Rapid burst kinetics in the hydrolysis of 4-nitrophenyl acetate by penicillin G acylase from Kluyvera citrophila. Effects of mutation F360V on rate constants for acylation and de-acylation

1996 ◽  
Vol 316 (2) ◽  
pp. 409-412 ◽  
Author(s):  
A Roa ◽  
M L Goble ◽  
J L García ◽  
C Acebal ◽  
R Virden
Keyword(s):  
Rate Constants ◽  
Acetyl Derivative ◽  
Penicillin G Acylase ◽  
Penicillin G ◽  
Side Chain ◽  
Wild Type ◽  
Wild Type Enzyme ◽  
Burst Kinetics ◽  
Kinetics Of ◽  
Hydrolysis Of

The kinetics of release of 4-nitrophenol were followed by stopped-flow spectrophotometry with two 4-nitrophenyl ester substrates of penicillin G acylase from Kluyvera citrophila. With the ester of acetic acid, but not of propionic acid, there was a pre-steady-state exponential phase, the kinetics of which were inhibited by phenylacetic acid (a product of hydrolysis of specific substrates) to the extent predicted from Ki values. This was interpreted as deriving from rapid formation (73 mM-1·s-1) and slow hydrolysis (0.76 s-1) of an acetyl derivative of the side chain of the catalytic-centre residue Ser-290. With the mutant F360V, which differs from the wild-type enzyme in its ability to hydrolyse adipyl-L-leucine and has a kcat for 4-nitrophenyl acetate one-twentieth that of the wild-type enzyme, the corresponding values for the rates of formation and hydrolysis of the acetyl-enzyme were 11.1 mM-1·s-1 and 0.051 s-1 respectively. The ratio of these rate constants was three times that for the wild-type enzyme, suggesting that the mutant is less impaired in the rate of formation of an acetyl-enzyme than in its subsequent hydrolysis.

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