Browse > Article

Purification and Characterization of Alkali-resistant Amylases from Pseudomonas sp.  

Lee, Jeong-Eun (Department of Food and Nutrition, Chonnam National University)
Jhon, Deok-Young (Department of Food and Nutrition, Chonnam National University)
Publication Information
Korean Journal of Food Science and Technology / v.40, no.1, 2008 , pp. 70-75 More about this Journal
Abstract
Two extracellular amylase isozymes were purified and characterized from alkalophilic Pseudomonas sp. KFCC 10818 for the production of maltooligosaccharides. The molecular weights of the homogeneous proteins were 50 kDa and 75 kDa, respectively. The 50 and 75 kDa amylases showed optimum temperatures at 35 and $40^{\circ}C$, respectively. The optimum pH of the enzymes ranged from pH 6-8, and the enzymes were resistant to an alkaline condition of pH 12. Via the enzyme's actions, the final products from maltooligosaccharides or soluble starch were maltose and maltotriose. Calcium was a potent activator of the 50 kDa amylase. Finally, the N-terminal amino acid sequences of the 50 and 75 kDa amylases were QTVPKTTFV and DTVPGNAFQ, respectively.
Keywords
amylase; purification; N-terminal sequencing; Pseudomonas sp.;
Citations & Related Records

Times Cited By SCOPUS : 0
연도 인용수 순위
  • Reference
1 Oishi M, Takahashi H, Maruo B. Intracellular ${\alpha}$-amylase in Bacillus subtilis. J. Bacteriol. 85: 246-247 (1963)
2 Takasaki Y. Purification and enzymatic properties of ${\beta}$-amylase and pullulanase from Bacillus cereus var. mycoides. Agric. Biol. Chem. 38: 1023-1029 (1976)
3 Shen GJ, Saha BC, Lee YE, Bhatnagar L, Zeikus JG. Purification and characterization of a novel thermostable ${\beta}$-amylase from Clostridium thermosulphurogenes. Biochem. J. 254: 838-840 (1988)
4 Gupta R, Gigras P, Mohapatra H, Goswami VK, Chauhan B. Microbial ${\alpha}$-amylases: A biotechnological perspective. Process Biochem. 38: 1599-616 (2003)   DOI   ScienceOn
5 Cheong KA, Tang SY, Cheong TK, Cha H, Kim JW, Park KH. Thermostable and alkalophilic maltogenic amylase of Bacillus thermoalkalophilus ET2 in monomer-dimer equilibrium. Biocatal. Biotransfor. 23: 79-87 (2005)   DOI   ScienceOn
6 Miller GL. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 31: 426-428 (1959)   DOI
7 Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the folin phenol reagent. J. Biol. Chem. 193: 265-275 (1951)
8 Laemmli UK. Cleavage of structural proteins during assembly of head of bacteriophage-T4. Nature 227: 680-685 (1970)   DOI   ScienceOn
9 Nielsen AD, Fuglsang CC, Westh P. Effect of calcium ions on the irreversible denaturation of a recombinant Bacillus halmapalus ${\alpha}$-amylase: a calorimetric investigation. Biochem. J. 373: 337-343 (2003)   DOI   ScienceOn
10 Oh KW, Kim MJ, Kim HY, Kim BY, Baik MY, Auh JH, Park CS. Enzymatic characterization of a maltogenic amylase from Lactobacillus gasseri ATCC 33323 expressed in Escherichia coli. FEMS Microbiol. Lett. 252: 175-181 (2005)   DOI   ScienceOn
11 Kato K, Sugimoto T, Amemura A, Harada T. Pseudomonas intracellular amylase with high activity on maltodextrins and cyclodextrins. Biochim. Biophys. Acta 391: 96-108 (1975)   DOI   ScienceOn
12 Simpson CL, Russell RRB. Intracellular ${\alpha}$-amylase of Streptococcus mutans. J. Bacteriol. 180: 4711-4717 (1998)
13 Hashim SO, Delgado OD, Martinez MA, Kaul RH, Mulaa FJ, Mattiasson B. Alkaline active maltohexaose-forming ${\alpha}$-amylase from Bacillus halodurans LBK 34. Enzyme Microb. Tech. 36: 139-146 (2005)   DOI   ScienceOn
14 Ballschmiter M, Futterer O, Liebl W. Identification and characterization of a novel intracellular alkaline ${\alpha}-$-amylase from the hyperthermophilic bacterium Thermotoga maritima MSB8. Appl. Environ. Microbiol. 72: 2206-2211 (2006)   DOI   ScienceOn
15 Rao JLUM, Satyanarayana T. Purification and characterization of a hyperthermostable and high maltogenic ${\alpha}$-amylase of an extreme thermophile Geobacillus thermoleovorans. Appl. Biochem. Biotech. 142: 179-193 (2007)   DOI   ScienceOn
16 Kang EJ, Kim ES, Lee JE, Jhon DY. Cloning, sequencing, characterization, and expression of a new ${\alpha}$-amylase isozyme gene (amy3) from Pseudomonas sp. Biotechnol. Lett. 23: 811-816 (2001)   DOI   ScienceOn
17 Bradford MM. Rapid and sensitive method for quantitation of microgram quantities of protein utilizing principle of protein-dye binding. Anal. Biochem. 72: 248-254 (1976)   DOI   ScienceOn
18 Igarashi K, Hatada Y, Hagihara H, Saeki K, Takaiwa M, Uemura T, Ara K, Ozaki K, Kawai S, Kobayashi T, Ito S. Enzymatic properties of a novel liquefying ${\alpha}$-amylase from an alkaliphilic Bacillus isolate and entire nucleotide and amino acid sequences. Appl. Environ. Microb. 64: 3282-3289 (1998)
19 Kim ES, Na HK, Jhon DY, Yoo OJ, Chun SB, Wui IS. Cloning, sequencing and expression of the amylase isozyme gene from Pseudomonas sp. KFCC 10818. Biotechnol. Lett. 18: 169-174 (1996)   DOI
20 Liu B, Wang YQ, Zhang XB. Characterization of a recombinant maltogenic amylase from deep sea thermophilic Bacillus sp. WPD616. Enzyme Microb. Tech. 39: 805-810 (2006)   DOI   ScienceOn
21 Lim WJ, Park SR, An CL, Lee JY, Hong SY, Shin EC, Kim EJ, Kim JO, Kim H, Yun HD. Cloning and characterization of a thermostable intracellular ${\alpha}$-amylase gene from the hyperthermophilic bacterium Thermotoga maritima MSB8. Res. Microbiol. 154: 681-687 (2003)   DOI   ScienceOn
22 Kim TU, Gu BG, Jeong JY, Byun SM, Shin YC. Purification and characterization of a maltotetraose-forming alkaline ${\alpha}-$-amylase from an alkalophilic Bacillus strain, GM8901. Appl. Environ. Microb. 61: 3105-3112 (1995)
23 Saxena RK, Dutt K, Agarwal L, Nayyar P. A highly thermostable and alkaline amylase from a Bacillus sp. PN5. Bioresource Technol. 98: 260-265 (2007)   DOI   ScienceOn
24 Na HK, Kim ES, Lee HB, Yoo OJ, Jhon DY. Cloning and nucleotide sequence of the ${\alpha}$-amylase gene from alkalophilic Pseudomonas sp. KFCC 10818. Mol. Cells 6: 203-208 (1996)
25 Das K, Doley R, Mukherjee AK. Purification and biochemical characterization of a thermostable, alkaliphilic, extracellular ${\alpha}$-amylase from Bacillus subtilis DM-03, a strain isolated from the traditional fermented food of India. Biotechnol. Appl. Bio. 40: 291-298 (2004)   DOI   ScienceOn
26 Whitehead TR, Cotta MA. Identification of intracellular amylase activity in Streptococcusbovis and Streptococcussalivarius. Curr. Microbiol. 30: 143-148 (1995)   DOI
27 Pandey A, Nigam P, Soccol CR, Soccol VT, Sing D, Mohan R. Advances in microbial amylases. Biotechnol. Appl. Bioc. 31: 135-152 (2000)   DOI   ScienceOn
28 Osman AB. Amylase in chicken intestine and pancreas. Comp. Biochem. Physiol. 73B: 571-574 (1982)
29 Chi ZM, Liu J, Zhang W. Trehalose accumulation from starch by Saccharomycopsis fibuligera sdu. Enzyme Microb. Tech. 28: 240-245 (2001)   DOI   ScienceOn
30 Bernhardsdotter ECMJ, Ng JD, Garriott OK, Pusey ML. Enzymic properties of an alkaline chelator-resistant ${\alpha}-$-amylase from an alkaliphilic Bacillus sp. isolate L1711. Process Biochem. 40: 2401-2408 (2005)   DOI   ScienceOn