Purification and characterization of a novel cold‐adapted phytase from Rhodotorula mucilaginosa strain JMUY14 isolated from Antarctic

Abstract

A yeast producing a cold-adapted phytase was isolated from Antarctic deep-sea sediment and identified as a Rhodotorula mucilaginosa strain JMUY14 of basidiomycetous yeasts. It was cultured in fermentation optimized by a response surface methodology based on the Box-Behnken design. The maximum activity of phytase reached 205.447 U ml(-1), which was close to the predicted value of 201.948 U ml(-1) and approximately 3.4 times higher than its initial activity. The extracellular phytase was purified by 15.2-fold to homogeneity with a specific activity of 31,635 U mg(-1) by (NH4 )2 SO4 precipitation, and a combination of DEAE Sepharose Fast Flow, SP Sepharose Fast Flow, and Sephadex G-100. The molecular weight of the purified enzyme was estimated to be 63 kDa and its pI was 4.33. Its optimal temperature and pH were 50 °C and 5.0, respectively. Its activity was 85% at 37 °C, and showed good stability at pH 3.0 ∼ 7.0. When compared with mesophilic counterparts, the phytase not only exhibited a higher activity during 20 ∼ 30 °C but also had a low Km (247 µM) and high kcat (1394 s(-1)). The phytase activity was slightly stimulated in the presence of Mg(2+), Fe(2+), Fe(3+), K(+), Na(+), Ca(2+), EDTA, and EGTA and moderately inhibited by Cu(2+), Zn(2+), Mn(2+), Ag(+), PMSF, SDS, and phenylgloxal hydrate. It was resistant to both pepsin and trypsin. Since the phytase produced by the R. mucilaginosa JMUY14 showed a high specific activity, good pH stability, strong protease resistance, and high activity at low temperature, it has great potential for feed applications, especially in aquaculture.

Production and Partial Characterization of an Extracellular Phytase Produced by <i>Muscodor</i> sp. under Submerged Fermentation

“…The maximum phytase activity (26.51 U/mg) was achieved at 144 h of fermentation (Figure 2(d)). This period for enzyme production was minor than that mentioned for production of phytase by Rhodotorula mucilaginosa JMUY [32]. On the other hand, it was higher than that observed for enzyme production by Rhizopus microsporus var.…”

Cold-adapted enzymes produced by fungi from terrestrial and marine Antarctic environments

“…Cold-adapted phytases might have considerable advantages in direct inclusion in feed for monogastric animals and use in aquaculture [79]. Pavlova et al [78] reported that Papiliotrema laurentii AL27 (formerly Cryptococcus laurentii) showed highest intracellular phytase activity at an optimal temperature of 40.0 C. In another study, Yu et al [79] reported the production of extracellular phytase by Rhodotorula mucilaginosa JMUY14, which showed resistance to both pepsin and trypsin degradation after purification.…”

“…The techniques used were diverse and yielded purification factors from 1.70-to 1568.00-fold and yield ranging from 0.65 to 86.20%. Enzymes secreted into the medium by Antarctic fungi are first concentrated by techniques, such as ultrafiltration [68,74,80,81,98,130], ammonium sulfate precipitation [63,64], ammonium sulfate and dialysis [79,85], or acetone precipitation [107]. After concentrating, the protein, purification can be carried out by a single or multiple steps of chromatography.…”

Characteristics of Recombinant Phytase (rSt-Phy) of the Thermophilic mold Sporotrichum thermophile and its applicability in dephytinizing foods

“…The application of phytase as the feed supplement has encouraged researchers to develop economical strategies to produce the enzyme in a cost-effective manner. Although a number of fungi, yeast, and bacteria are known to produce phytases [3,4,28,29], none of them produce adequate enough to produce it on a large scale. The cloning and over expression of the enzymes is an approach that permits enzyme manufacture in a cost-effective manner.…”