Streptococcus infantarius 25124 isolated from pozol produces a high molecular weight amylopullulanase, a key enzyme for niche colonization

Abstract

Pozol is a beverage made with maize dough that is prepared after boiling the kernels in limewater, causing a decrease in soluble sugars, with starch being the main fermentable carbohydrate in the dough. Previously, Streptococcus infantarius ssp. infantarius 25124 (Sii-25124) was identified as the most amylolytic bacteria isolated in this product. Analysis of Sii-25124 amylolytic enzymes revealed two amylases, a cytoplasmic α-amylase of 55.7 kDa and an extracellular amylopullulanase of 246.3 kDa, with two catalytic domains, one typical of an α-amylase and another typical of a pullulanase/glycogen debranching enzyme. Characterization of the joint activity of both enzymes using Sii-25124 cell lysate supernatant demonstrated stability between 30 °C and 45°C, and pH stability in a range between 6.8 and 8.0. The joint activity of Sii-25124 amylases showed a fast production of reducing sugars when starch was used as the substrate. In contrast, reducing sugar production from amylopectin was lower, but it steadily increased throughout the reaction time. The amylopullulanase produced by Sii-25124 hydrolyzes the starch in the dough to produce low molecular weight oligosaccharides, which may be transported into Sii-25124 cells, so that intracellular α-amylase hydrolyzes them to mono- and disaccharides. Amylopullulanase production by Sii-25124 could be an example of a specialized enzyme that successfully dominates starchy food fermentation.

An Alkalothermophilic Amylopullulanase from the Yeast Clavispora lusitaniae ABS7: Purification, Characterization and Potential Application in Laundry Detergent

“…The differential behavior of the activities of α-amylase and pullulanase regarding certain metal ions (such as Ca 2+ , Hg 2+ , and Mg 2+ ) may be due to the presence of two different active sites, one for the α-amylase and the other for the pullulanase [63]. Compared to the control, the effect of divalent ions on amylolytic activity showed that the presence of Mg 2+ increased amylolytic activity by 146%, while Mn 2+ , Fe 3+ , Ca 2+ , and Na + increased amylolytic activity to 141%, 116%, 112%, and 111%, respectively [41]. It is known that most amylases are considered metalloenzymes, which are enzymes that require metal ions (usually Ca 2+ ) to maintain their stable native structure.…”

“…A previous work [41] reported the optimal temperature and pH of amylopullulanase from Streptococcus infantarius ssp. as 37 • C and 6.8, respectively.…”

Analysing the dynamics of the bacterial community in pozol, a Mexican fermented corn dough

“…Consequently, the core microbiota responsible for corn fermentation is expected to have this metabolic capacity, as has already been shown in S. infantarius , isolated from pozol [14, 43]. On the other hand, while the alkaline thermal treatment associated with nixtamal removes most soluble sugars, it also predigests complex insoluble carbohydrates from the plant cell wall, increasing the availability of arabinoxylan, the main component of the hemicellulose present in fermented maize dough [44, 45].…”

Metaproteomic Insights Into the Microbial Community in Pozol

“…Characterization of the most amylolytic LAB species, Streptococcus infantarius subsp. infantarius , produced two amylases, an amylopululanase with debranching capacity and an α-amylase ( 9 , 57 ).…”