Syed Shahid Ali

Improvement of penicillin G acylase expression in Escherichia coli through UV induced mutations.

We used ultraviolet (UV) radiation to induce mutation in three locally isolated strains of Escherichia coli. Different dilutions of bacterial cultures were exposed to UV lamp of 254 nm wavelength for different time intervals at varied distances ranging from 5 to 210 sec and 5 to 100 cm. Viable colonies were screened for mutants with an increased production of penicillin G acylase (PGA) and a reduced production of β-lactamase, which are the desired properties of PGA producing industrial strains. A survival curve was made to get optimum exposure time and distance. The survival percentage for each exposure period was calculated and 1-5% survival was found useful for obtaining mutants with desired change. Screening for PGA and β-lactamase constitutive and/or deficient mutants was made by Serratia marcescens overlay test. A total of 100 survivors were selected of which 49% expressed PGA activity higher than the parent strain. Frequency of β-lactamase constitutive and deficient mutants was 48 and 52%, respectively. The best hyper-producing mutant (BDCS-N-M74), with almost negligible expression of β-lactamase, exhibited three-fold (22.5 mg 6-APA h-1 mg-1 wet cells) increase in PGA activity compared with that in the parent strain (6.7 mg 6-APA h-1 mg-1 wet cells). The results indicated the successful induction of UV mediated mutation in E. coli for PGA hyper-producing mutants lacking β-lactamase activity.

Mutagenic effect of acridine orange on the expression of penicillin G acylase and β-lactamase in Escherichia coli

Aims:  The present work aimed to improve the production of penicillin G acylase (PGA) and reduce the β‐lactamase activity through acridine orange (AO) induced mutation in Escherichia coli.

Relative Activity of Various Esterases in Six Pakistani Strains of the Lesser Grain Borer, Rhyzopertha dominica (Fabricius)

Acetylcholinesterase (AChE), arylesterase (AE), carboxylesterase (CE), and cholinesterase (ChE) activities were determined in six adult populations of the lesser grain borer, Rhyzopertha dominica (Fabricius) (Coleoptera: Bostrichidae) collected from grain warehouses in various areas of Punjab, namely Lahore (L), Sialkot (S), Wazirabad (W), Multan (M), Chichawatni (C), and Karachi (K). The multiple forms of esterases were resolved by polyacrylamide gel electrophoresis (PAGE) and activities determined biochemically, using a spectrophotometer. All the strains had the same AChE activity except Sialkot strain, which had 46% more activity than the susceptible K strain. Similarly K, S, W, L, and C strains exhibited almost the same AE level, whereas the M strain showed 29% lower activity than K strain. The CE activity of L (352%), S (198%) and C (214%) strains and ChE activity of L (178%) and C (317%) strains were significantly higher than that of K strain. The total esterases of L, S and C strains showed respectively 152%, 63% and 73% higher activity than the susceptible K strain. The PAGE pattern of various esterases coincided with the biochemical analysis. The variable thickness of the bands in the gel indicated relative esterase induction that could be correlated with the development of resistance in R. dominica.RésuméLes activités acétylcholinestérasique, arylestérasique, carboxylestérasique, et cholinestérasique ont été déterminées dans six populations d’adultes du foreur de grain, Rliyzopertlia dominica (F.), collectés à partir des entrepôts de grain dans diverses régions du Punjab, à savoir Lahore (L), Sialkot (S), Wazitabad (W), Multan (M), Chkhawatni (C), et Karachi (K). Diverses formes d’estérases ont été résolues par électrophorèse à gel de polyacrylamide et les activités biochimiquement déterminées par spectrophotomètrie. Toutes les souches avaient la mê;me activité acétylcholinestérasique, excepté la souche S, qui a eu 46% de plus d’activité que la souche K sensible. Les souches K, S, W, L, et C ont montré presque le même niveau d’arylestérase, tandis que la souche M avait 29% de moins d’activité que la souche K. L’activité carboxylestérasique des souches L (352%), S (198%) et C (214%) et l’activité cholinestérasique des souches L (178%) et C (317%) étaient sensiblement plus élevées que celles de la souche K. Toutes les estérases des souches L, S et C ont respectivement montré 152%, 63% et 73% d’activité plus élevée que la souche K sensible. La configuration de l’électrophorèse à gel de polyacrylamide de diverses estérases a coïncidé avec leurs analyses biochimiques. La variabilité de l’épaisseur des bandes dans le gel a suggeré l’induction relative d’estérase qui pourrait ê;tre corrélée avec le développement de la résistance au sein des pupulations de R. dominica.