Overexpression of alginate lyase of Pseudoalteromonas elyakovii in Escherichia coli, purification, and characterization of the recombinant alginate lyase

Abstract

The alyPEEC gene encoding alginate lyase from marine bacterium Pseudoalteromonas elyakovii IAM 14594 was subcloned into pBAD24 with arabinose promoter and sequenced, and overexpressed in TOP10 strain of E. coli after arabinose induction. Expression levels of alyPEEC gene in E. coli cells were over 39.6-fold higher than those in P. elyakovii IAM 14594 cells. The molecular mass of purified alginate lyase from the engineered E. coli cells was estimated to be 32.0 kDa. Optimum pH and temperature of the alginate lyase activity were 7.0 and 30°C, respectively. The enzyme was unstable on heating and in acidic and alkaline solution. The enzyme activity was stimulated by the MgCl 2 , NaCl, KCl, CaCl 2 , BaCl 2 and MnCl 2 , but was inhibited by the addition of 1.0 mM of EGTA, EDTA, SDS, ZnSO 4 , AgNO 3 , and CoCl 2 . All the alginate, polyM and polyG could be converted into oligosaccharides with more than tetrasaccharides by the purified recombinant alginate lyase, suggesting that the recombinant alginate lyase produced by the engineered E. coli has highly potential application in seaweed genetics, food and pharmaceutical industries.

Characterization of a Novel Alginate Lyase from Marine Bacterium Vibrio furnissii H1

“…The reaction mixture containing 1 mL of 1.0% sodium alginate and 1 mL of enzyme was incubated at 30 °C for 24 h. A 5 μL aliquot of the reaction product was subjected to TLC using a solvent system of 1-butanol: acetic acid: water (3:2:2, v / v ). The plate was then sprayed with 10% ( v / v ) sulfuric acid in ethanol and heated at 120 °C for 5 min [ 41 ].…”

Expression and Characterization of a New PolyG-Specific Alginate Lyase From Marine Bacterium Microbulbifer sp. Q7

“…Alginate lyase present in diverse organisms (Singh et al, 2011 ; Wang et al, 2013 ), degrades alginate by β-elimination and produces unsaturated oligosaccharides with double bonds at the non-reducing end (Ma et al, 2008 ). Alginate degradation products have been developed for broad applications in agricultural, health product and medical industries, since they exhibit biological activities including immunomodulation, anti-tumor, antioxidant, and plant growth-promoting activities (Yokose et al, 2009 ; Tusi et al, 2011 ; Zhou et al, 2015 ).…”

A novel Bacillus sp. isolated from rotten seaweed: Identification and characterization alginate lyase its produced

“…(Iwamoto et al 2001), Corynebacterium sp. (Matsubara et al1998), Pseudoalteromonas elyakovii (Ma et al 2008), Pseudomonas syringae (Preston et al 2000), and Streptomyces sp. (Cao et al 2007;Kim et al 2009).…”