Biochemical Characterization of Kluyveromyces lactis Adenine Deaminase and Guanine Deaminase and Their Potential Application in Lowering Purine Content in Beer

Abstract

Excess amounts of uric acid in humans leads to hyperuricemia, which is a biochemical precursor of gout and is also associated with various other disorders. Gout is termed as crystallization of uric acid, predominantly within joints. The burden of hyperuricemia and gout has increased worldwide due to lifestyle changes, obesity, and consumption of purine-rich foods, fructose-containing drinks, and alcoholic beverages. Some of the therapies available to cure gout are associated with unwanted side-effects and antigenicity. We propose an attractive and safe strategy to reduce purine content in beverages using enzymatic application of purine degrading enzymes such as adenine deaminase (ADA) and guanine deaminase (GDA) that convert adenine and guanine into hypoxanthine and xanthine, respectively. We cloned, expressed, purified, and biochemically characterized both adenine deaminase (ADA) and guanine deaminase (GDA) enzymes that play important roles in the purine degradation pathway of Kluyveromyces lactis, and demonstrate their application in lowering purine content in a beverage. The popular beverage beer has been selected as an experimental sample as it confers higher risks of hyperuricemia and gout. Quantification of purine content in 16 different beers from the Indian market showed varying concentrations of different purines. Enzymatic treatment of beer samples with ADA and GDA showed a reduction of adenine and guanine content, respectively. These enzymes in combination with other purine degrading enzymes showed marked reduction in purine content in beer samples. Both enzymes can work at 5.0–8.0 pH range and retain >50% activity at 40°C, making them good candidates for industrial applications.

The potential of probiotics in the amelioration of hyperuricemia

“…57 A recent study found that several probiotics, especially those produced by the Lactobacillus genus, can degrade inosine and guanosine to lower the levels of metabolites such as ribose and base. 58 Since the human body cannot synthesize vitamins, studies have shown that the purine metabolic pathway changes after the intake of specific probiotics, and newly colonized strains in the intestine take up ribose or nucleotides as raw materials to synthesize alkaloids, such as thiamine, riboflavin, and folic acid. 59,60 With the reduction in these intermediates, uric acid synthesis is slowed, and the development of probiotics that effectively degrade purine compounds into metabolites other than uric acid is a potential method for preventing and/or treating hyperuricemia.…”

Towards the Use of Adsorption Methods for the Removal of Purines from Beer

“…These findings suggest the applicability of KlacPNP and respective variants on the crucial steps that contribute to decrease the purine contents in beer. Moreover, Mahor et al [65] used adenine deaminase and guanine deaminase, involved in the conversion of adenine to hypoxanthine and guanine to xanthine, respectively, of Kluyveromyces lactis, to reduce the purines content in beer. A reduction on adenine contents of 66-67% and a decrease of guanine concentration from 68.8 µmol L −1 to residual values were found.…”

“…(1) uric acid overproduction and/or excessive intake, (2) impair uric acid excretion and (3) a combination of both [53,55,65]. Mechanisms should exist to ensure the efficient disposal of uric acid and, since humans are incapable to metabolize it, excretion processes play a crucial role in maintaining homeostasis [30].…”

Carboxylated Cellulose as a Soft Template Combined with PEDOT Derivatives in [BMIM]Cl: A Competent Biosensor for Detection of Guanine and Uric Acid in the Blood

“…Poly (3,4-ethylendioxythiophene) (PEDOT) is a conductive polymer that has been proposed as an alternative to traditional polymers used as the electroactive component in biosensors. 9 In recent years, the application of PEDOT for the detection of heavy metal ions and biological small molecules has also become a research hotspot.…”

“…Moreover, due to the similar structures of G and UA, they coexist in the human body, are present in the same metabolic pathways, and cause various detection problems. 3 Therefore, the development of a sensitive and selective method for the simultaneous determination of G and UA is highly desirable in the biomedical chemistry field.…”