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Production of diamino propionic acid ammonia lyase by a new strain of Salmonella typhimurium PU011.

Rupesh KR, PremKumar PL, Shiva Kumar VV, Jayachandran SS - BMC Microbiol. (2002)

Bottom Line: Following ammonium sulphate precipitation and passing through Sephadex G100, CM-Sephadex and DEAE-Sephacel for crude enzyme extract preparation, about 68-fold enzyme purity was obtained.The purified enzyme gave maximum activity at pH 8.0 and was stable up to 45 degrees C.The Km value for the substrate was found to be 0.685 mM, calculated from a Line Weaver Burk plot.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biotechnology, Pondicherry University, Kalapet, Pondicherry 605014, India. krrupesh@yahoo.com

ABSTRACT

Background: Seeds of the legume plant Lathyrus sativus, which is grown in arid and semi arid tropical regions, contain Diamino Propionic acid (DAP). DAP is a neurotoxin, which, when consumed, causes a disease called Lathyrism. Lathryrism may manifest as Neurolathyrism or Osteolathyrism, in which the nervous system, and bone formation respectively, are affected. DAP ammonia lyase is produced by a few microorganisms such as Salmonella typhi, Salmonella typhimurium and Pseudomonas, and is capable of detoxifying DAP.

Results: S. typhimurium PU011, a non-virulent bacterial strain isolated in our lab, was found to produce DAP ammonia lyase enzyme when grown in minimal medium containing DAP. There was a direct correlation between biomass yield and enzyme activity, until 16 h post inoculation in minimal medium containing DAP. Following ammonium sulphate precipitation and passing through Sephadex G100, CM-Sephadex and DEAE-Sephacel for crude enzyme extract preparation, about 68-fold enzyme purity was obtained. The purified enzyme gave maximum activity at pH 8.0 and was stable up to 45 degrees C. The Km value for the substrate was found to be 0.685 mM, calculated from a Line Weaver Burk plot.

Conclusion: A new bacterial strain, S.typhimurium PU 011, which is capable of producing DAP ammonia lyase, was isolated.

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Biomass and DAP ammonia lyase production in Salmonella typhimurium PU 011. The culture was incubated on a rotary shaker at room temperature. The biomass was estimated gravimetrically and expressed as mg/ml. The crude enzyme activity of the culture drawn at different intervals of growth was measured spectrophotometrically and expressed as (U/mg) × 10-3.
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Figure 1: Biomass and DAP ammonia lyase production in Salmonella typhimurium PU 011. The culture was incubated on a rotary shaker at room temperature. The biomass was estimated gravimetrically and expressed as mg/ml. The crude enzyme activity of the culture drawn at different intervals of growth was measured spectrophotometrically and expressed as (U/mg) × 10-3.

Mentions: Vijayalakshmi (1975) [6] and Nagasawa (1988) [7] have identified Pseudomonad and Salmonella typhimurium respectively capable of producing DAP ammonia lyase that degrades the neurotoxin present in L. sativus seeds. We have accidentally stumbled upon a non-virulent bacterium S. typhimurium PU011 which was capable of producing DAP ammonia lyase. The kinetics of production of biomass and DAP ammonia lyase activity by this organism is shown in Fig 1. A continuous increase in biomass and specific activity of enzyme with increase in incubation period was observed, the maximum reaching at 16 h post inoculation. Interestingly, a direct correlation between biomass yield and enzyme activity up to 20 h after inoculation was observed. Nagasawa et al. (1988)[7] have also observed a similar pattern of growth and enzyme activity in S. typhimurium.


Production of diamino propionic acid ammonia lyase by a new strain of Salmonella typhimurium PU011.

Rupesh KR, PremKumar PL, Shiva Kumar VV, Jayachandran SS - BMC Microbiol. (2002)

Biomass and DAP ammonia lyase production in Salmonella typhimurium PU 011. The culture was incubated on a rotary shaker at room temperature. The biomass was estimated gravimetrically and expressed as mg/ml. The crude enzyme activity of the culture drawn at different intervals of growth was measured spectrophotometrically and expressed as (U/mg) × 10-3.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC101404&req=5

Figure 1: Biomass and DAP ammonia lyase production in Salmonella typhimurium PU 011. The culture was incubated on a rotary shaker at room temperature. The biomass was estimated gravimetrically and expressed as mg/ml. The crude enzyme activity of the culture drawn at different intervals of growth was measured spectrophotometrically and expressed as (U/mg) × 10-3.
Mentions: Vijayalakshmi (1975) [6] and Nagasawa (1988) [7] have identified Pseudomonad and Salmonella typhimurium respectively capable of producing DAP ammonia lyase that degrades the neurotoxin present in L. sativus seeds. We have accidentally stumbled upon a non-virulent bacterium S. typhimurium PU011 which was capable of producing DAP ammonia lyase. The kinetics of production of biomass and DAP ammonia lyase activity by this organism is shown in Fig 1. A continuous increase in biomass and specific activity of enzyme with increase in incubation period was observed, the maximum reaching at 16 h post inoculation. Interestingly, a direct correlation between biomass yield and enzyme activity up to 20 h after inoculation was observed. Nagasawa et al. (1988)[7] have also observed a similar pattern of growth and enzyme activity in S. typhimurium.

Bottom Line: Following ammonium sulphate precipitation and passing through Sephadex G100, CM-Sephadex and DEAE-Sephacel for crude enzyme extract preparation, about 68-fold enzyme purity was obtained.The purified enzyme gave maximum activity at pH 8.0 and was stable up to 45 degrees C.The Km value for the substrate was found to be 0.685 mM, calculated from a Line Weaver Burk plot.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biotechnology, Pondicherry University, Kalapet, Pondicherry 605014, India. krrupesh@yahoo.com

ABSTRACT

Background: Seeds of the legume plant Lathyrus sativus, which is grown in arid and semi arid tropical regions, contain Diamino Propionic acid (DAP). DAP is a neurotoxin, which, when consumed, causes a disease called Lathyrism. Lathryrism may manifest as Neurolathyrism or Osteolathyrism, in which the nervous system, and bone formation respectively, are affected. DAP ammonia lyase is produced by a few microorganisms such as Salmonella typhi, Salmonella typhimurium and Pseudomonas, and is capable of detoxifying DAP.

Results: S. typhimurium PU011, a non-virulent bacterial strain isolated in our lab, was found to produce DAP ammonia lyase enzyme when grown in minimal medium containing DAP. There was a direct correlation between biomass yield and enzyme activity, until 16 h post inoculation in minimal medium containing DAP. Following ammonium sulphate precipitation and passing through Sephadex G100, CM-Sephadex and DEAE-Sephacel for crude enzyme extract preparation, about 68-fold enzyme purity was obtained. The purified enzyme gave maximum activity at pH 8.0 and was stable up to 45 degrees C. The Km value for the substrate was found to be 0.685 mM, calculated from a Line Weaver Burk plot.

Conclusion: A new bacterial strain, S.typhimurium PU 011, which is capable of producing DAP ammonia lyase, was isolated.

Show MeSH
Related in: MedlinePlus