Production of L-asparaginase by filamentous fungi

Abstract

L-asparaginase production was investigated in the filamentous fungi Aspergillus tamarii and Aspergillus terreus.The fungi were cultivated in medium containing different nitrogen sources. A. terreus showed the highest L-asparaginase (activity) production level (58 U/L) when cultivated in a 2% proline medium. Both fungi presented the lowest level of L-asparaginase production in the presence of glutamine and urea as nitrogen sources. These results suggest that L-asparaginase production by of filamentous fungi is under nitrogen regulation.Key words: filamentous fungi -production of L-asparaginase -Aspergillus tamarii -Aspergillus terreus Asparaginase (L-asparagine amido hydrolase, E.C. 3.5.1.1) is an anti-neoplastic agent, used in the lymphoblastic leukaemia chemotherapy. Neoplastic cells cannot synthetase L-asparagine due the absence of L-asparagine synthetase (Keating et al. 1993). For this reason the commonest therapeutic practice is to inject intravenously free enzyme in order to decrease the blood concentration of Lasparagine affecting selectively the neoplastic cells (Mitchell et al. 1994). However, L-asparaginase from bacterial origin can cause hypersensitivity in the long-term used, leading to allergic reactions and anaphylaxis (Reynolds & Taylor 1993). The search for other asparaginase sources, like eucaryotic microorganisms, can lead to an enzyme with less adverse effects. The importance of microorganisms as L-asparaginase sources has been focused since the time it was obtained from Escherichia coli and its antineoplastic activity demonstrated in guinea pig serum (Broome 1961, Mashburn & Wriston 1964, Roberts et al. 1966, Schawartz et al. 1966. The E. coli and Erwinia enzymes were isolated, purified, and experimentally used as an anti-leukaemic agent in human patients (Clavell et al. 1986, Story et al. 1993. It demonstrated high potential against children's acute lymphoblastic leukaemia (Hill et al. 1967, Oettgen et al. 1967. Several research groups have studied asparaginase production and purification in attempt to minimize impurities that produce allergenic reations , Boss 1997, Gallagher et al. 1999.It has been observed that eukaryote microorganisms like yeast and filamentous fungi have a potential for asparaginase production (Wade et al. 1971, Wiame et al.1985, Pinheiro et al. 2001 asparaginase (De-Angeli et al. 1970, Arima et al. 1972, Curran et al. 1985.L-asparaginase belongs to an amidase group that produces aspartic acid and ammonia by asparagine hydrolysis (Wriston & Yellin 1973, Capizzi et al. 1984. Here we present a simple and rapid assay method for the detection of asparaginase from different strains of Aspergillum, Penicillium, and Fusarium genera of fungi using the Gulati et al. (1997) method. After an initial screening, strains presenting high asparaginase activity were cultivated in submerged fermentation in order to determine the production of this enzyme in the presence of different nitrogen sources. Moreover the question whether asparaginase from filamentous fungi is nitrogen regulated a...

l-Asparaginase Production by Moderate Halophilic Bacteria Isolated from Maharloo Salt Lake

“…For example, Mycobacterium tuberculosis produces two kinds of L-asparaginase in the cell, but only one of them is effective against animal tumors. E. coli L-asparaginase I and II were produced simultaneously in the cells, but only the latter showed antitumor activity [3]. This different activity is due to differences in a number of properties, such as pH activity profile, sensitivity to thermal inactivation, clearance rate (half life in serum) and most significantly their affinity for L-asparagine [17,18].…”

“…For this reason, the commonest therapeutic practice is to inject the enzyme intravenously in order to decrease the blood concentration of L-asparagine, selectively affecting the neoplastic cells [1]. Since the observation that L-asparaginase from Escherichia coli has an antitumor activity similar to that of guinea pig serum, there has been considerable interest in asparaginase from various sources specially microorganisms [3]. Although other microorganisms such as Aerobacter, Bacillus, Erwinia, Pseudomonas, Serratia, Xanthomonas, Photobacterium [1], Streptomyces [4], Proteus [5], Vibrio [6] and Aspergillus [3] have a potential for asparaginase production, just the purified enzyme from E. coli and Erwinia sp.…”

Purification and Biochemical Characterization of Native and Pegylated Form of L-Asparaginase from <i>Aspergillus terreus</i> and Evaluation of Its Antiproliferative Activity

“…Enzyme production was optimized in Czapek Dox's modified liquid medium in two steps: the pre-fermentation medium containing 0.2% (w/v) glucose, 2% (w/v) L-proline, 0.2% (w/v) NH 4 7 spores·mL −1 and incubated at 120 rpm for 17 h at 30˚C. The culture was filtrated; the mycelium was collected and inoculated in the fermentative medium, which was similar to the medium used in previous step except for the absence of NH 4 NO 3 .…”

“…Over the years, several bacterial L-asparaginases have been reported and only a few reports about L-asparaginase produced by filamentous fungi have been made. Among these reports are the L-asparaginase production by Aspergillus tamari, A. terreus [4], A. niger [5], A. nidulans [6], and in some yeast, but their antiproliferative activeties were not analyzed. Currently, there are three asparaginases preparations available for therapy, two of them are native and produced by the bacteria Escherichia coli and Erwinia chrysanthemi.…”

Influence of Glucose and L-Asparagine Concentrations on L-Asparaginase Production by Endophytic Fungi

“…As an anticancer agent, L-asparaginase removes the L-asparagine in the blood, depriving tumor cells of L-asparagine, leading towards incapacitated cell division. L-asparaginase is an enzyme produced by animals, plants, yeasts and microorganisms (Sarquis et al, 2004). Among these, bacteria-produced Lasparaginase is the most extensively explored, due to their cost-effective nature (Theantana et al, 2009).…”