Purification and characterization of chitinases and chitosanases from a new species strain Pseudomonas sp. TKU015 using shrimp shells as a substrate.
A chitinase (CHT1) and a chitosanase (CHS1) were purified from the culture supernatant of Pseudomonas sp. TKU015 with shrimp shell wastes as the sole carbon and nitrogen source. The optimized conditions of this new species strain (Gen Bank Accession Number EU103629) for the production of chitinases were found to be when the culture was shaken at 30 degrees C for 3 days in 100 mL of medium (pH 8) containing 0.5% shrimp shell powder (SSP) (w/v), 0.1% K2HPO4, and 0.05% MgSO(4).7H2O. The molecular weights of CHT1 and CHS1 determined by SDS-PAGE were approximately 68 kDa and 30 kDa, respectively. The optimum pH, optimum temperature, pH stability, and the thermal stability of CHT1 and CHS1 were pH 6, 50 degrees C, pH 5-7, <50 degrees C and pH 4, 50 degrees C, pH 3-9, <50 degrees C, respectively. CHT1 was inhibited completely by Mn2+ and Fe2+, and CHS1 was inhibited by Mn2+, Cu2+, and PMSF. CHT1 was only specific to chitin substrates, whereas the relative activity of CHS1 increased when the degree of deacetylation of soluble chitosan increased.
Wang
San-Lang
SL
Graduate Institute of Life Sciences, Tamkang University, 151 Ying-Chuan Road, Tamsui 251, Taiwan. sabulo@mail.tku.edu.tw
Chen
Shin-Jen
SJ
Wang
Chuan-Lu
CL
D000107:Acetylation; D000818:Animals; D002244:Carbon / Q000378:metabolism*; D002686:Chitin / Q000494:pharmacology; D002688:Chitinases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D048271:Chitosan / Q000378:metabolism; D003470:Culture Media; D033364:Decapoda; D005296:Ferrous Compounds / Q000494:pharmacology; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D017895:Manganese Compounds / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009584:Nitrogen / Q000378:metabolism*; D011549:Pseudomonas / Q000201:enzymology*; D012685:Sepharose / Q000031:analogs & derivatives / Q000737:chemistry; D013379:Substrate Specificity; D013696:Temperature
D003470:Culture Media; D005296:Ferrous Compounds; D017895:Manganese Compounds; D002686:Chitin; C043982:DEAE-Sepharose CL-6B; C020927:phenyl-sepharose; D002244:Carbon; D012685:Sepharose; D048271:Chitosan; D006026:Glycoside Hydrolases; C019868:chitosanase; D002688:Chitinases; D009584:Nitrogen