RESUMO
Exposure to ultraviolet (UV) radiation is one of the major factors that causes skin aging, erythema, sunburns, and skin cancer. This study aimed to select probiotic bacterial isolates able to produce high yield of hyaluronic acid (HA) to be employed for skin photoprotection and other possible biological applications. The selected isolates K11 and St3 were able to produce the highest yields of HA 4.8 and 4.4 mg/ml, respectively. Both isolates were identified as Enterococcus durans strain K11 and Lactiplantibacillus plantarum strain St3 using 16S rRNA gene sequencing. The antioxidant activity of HA produced by E. durans strain K11 and L. plantarum strain St3 was (65.4 0.2%) and (66.6 0.1%), respectively. The viability of UVB-irradiated keratinocytes pre-treated with HA produced by E. durans strain K11 and L. plantarum strain St3 was 91.3 and 91.4%, respectively, compared with the control. While the viability of UVB-irradiated keratinocytes post-treated with HA produced by E. durans strain K11 and L. plantarum strain St3 was 86 and 88.5%, respectively. To the best of our knowledge, this is the first recordation of HA production by Enterococcus durans and Lactiplantibacillus plantarum which revealed a significant radioprotection of the human keratinocytes against UVB radiation.
Assuntos
Enterococcus , Ácido Hialurônico , Humanos , RNA Ribossômico 16S , PeleRESUMO
Background: Cyclodextrin glucanotransferase (CGTase) is one of the most industrially important enzymes used in the commercial production of cyclodextrins (CDs). Alkaliphilic bacteria have attracted much interest in the last few decades because of their ability to produce extracellular enzymes that are active and stable at high pH values. Here, we report the isolation of a new CGTase from alkaliphilic bacteria collected from Egyptian soda lakes and describe the purification and biochemical characterization of this CGTase. Results: Screening for CGTase-producing alkaliphilic bacteria from sediment and water samples collected from Egyptian soda lakes located in the Wadi Natrun valley resulted in the isolation of a potent CGTase-producing alkaliphilic bacterial strain, designated NRC-WN. Strain NRC-WN was belonging to genus Amplibacullus by 16S rDNA sequence analysis (similarity: ca. 98%). Among the tested nitrogen and carbon sources, peptone (0.15%, w/v) and soluble starch (0.4%, w/v) allowed maximal CGTase production by Amphibacillus sp. NRC-WN. CGTase was successfully purified from Amphibacillus sp. NRC-WN up to 159.7-fold through a combination of starch adsorption and anion exchange chromatography, resulting in a yield of 84.7%. SDS-PAGE analysis indicated that the enzyme was purified to homogeneity and revealed an estimated molecular mass of 36 kDa, which makes it one of the smallest CGTases reported in the literature. The purified enzyme exhibited maximum activity at 50ºC and was stable up to 70ºC, retaining 93% of its initial activity after treatment for 1 hr. Furthermore, Ca2+ ions (10 mM) significantly enhanced the thermal stability of the CGTase. The purified enzyme was active and stable over a wide pH range, showing maximal activity at pH 9.5. The enzyme was significantly stimulated by Zn2+, Ca2+ and Co2+ but was completely inhibited in the presence of Fe3+ and mercaptoethanol. The Km and Vmax values of the purified CGTase were estimated to be 0.0434 mg/ml and 3,333.3 mg β-CD/ml/min, respectively. β-CD was the predominant product of starch degradation by the Amphibacillus sp. NRC-WN CGTase, followed by α-and γ-CDs. Conclusions: A new low molecular mass alkaline CGTase was purified from a newly identified alkaliphilic Amphibacillus sp. NRC-WN isolate from the Egyptian soda lakes. The enzyme showed promising thermal and pH stability and a high affinity toward starch as a natural substrate.
