Killer toxin of Pichia anomala NCYC 432;: purification, characterization and its exo-β-1,3-glucanase activity

Pichia anomala NCYC 432 secretes a killer toxin which is inhibitory to a variety of yeasts including pathogenic Candida spp. The killer toxin in the culture supernatant was concentrated by ultratiltration and purified to homogenity by two successive gel filtration chromatographies with a TSK G2000SW column. Biochemical characterization of the toxin showed that it is a glycosylated protein with a molecular mass of 47 kDa and pI values of 3.4 and 3.7. The toxin showed high stability at pH values between 3 and 5.5 and up to 37 degrees C. Its N-terminal amino acid sequencing yielded the sequence GDYWDYQNDKIR which is 100% identical with that of mature exo-beta-1,3-glucanase (accession no. AJ222862) of P. anomala strain K. The toxin displayed high activity against laminarin thus showing a beta-glucanase activity. The Michaelis constants K-m and V-max for laminarin hydrolysis were 0.3 mg ml(-1) and 350 mu mol min(-1) mg(-1). (c) 2005 Elsevier Inc. All rights reserved.