title: Preservation of high phenylalanine ammonia lyase activities in roots of Japanese Striped corn: a potential oral therapeutic to treat phenylketonuria.
abstract: Phenylketonuria (PKU) is an inherited metabolic disorder caused by deficient phenylalanine hydroxylase (PAH) activity, the enzyme responsible for the disposal of excess amounts of the essential amino acid phenylalanine (Phe). Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) has potential to serve as an enzyme substitution therapy for this human genetic disease. Using 7-day-old Japanese Striped corn seedlings (Japonica Striped maize, Zea mays L. cv. japonica) that contain high activities of PAL, we investigated a number of methods to preserve the roots as an intact food and for long-term storage. The cryoprotectant effects of maple syrup and other edible sugars (mono- and oligosaccharides) were evaluated. Following thawing, the preserved roots were then examined to determine whether the rigid plant cell walls could protect the PAL enzyme from proteolysis during simulated (in vitro) digestion comprised of gastric and intestinal phases. While several treatments led to retention of PAL activity during freezing, upon thawing and in vitro digestion, root tissues that had been previously frozen in the presence of maple syrup exhibited the highest residual PAL activities (∼50% of the initial enzyme activity), in marked contrast to all of the treatments using other edible sugars. The structural integrity of the root cells, and the stability of the functional PAL tetramer were also preserved with the maple syrup protocol. These results have significance for the formulation of oral enzyme/protein therapeutics. When plant tissues are adequately preserved, the rigid cell walls constitute a protective barrier even under harsh (e.g. gastrointestinal-like) conditions.
authors.affiliation: Department of Biological Sciences, 8888 University Dr., Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada. Electronic address: artlopezvillalobos@hotmail.com.
authors.forename: Arturo
authors.initials: A
authors.lastname: López-Villalobos
authors.affiliation: Department of Biological Sciences, 8888 University Dr., Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada. Electronic address: jlucker@gmail.com.
authors.forename: Joost
authors.initials: J
authors.lastname: Lücker
authors.affiliation: Department of Biological Sciences, University of Calgary, 2500 University Drive N.W., Calgary, Alberta T2N 1N4, Canada. Electronic address: aalopezq@ucalgary.ca.
authors.forename: Ana Angela
authors.initials: AA
authors.lastname: López-Quiróz
authors.affiliation: Department of Biological Sciences, University of Calgary, 2500 University Drive N.W., Calgary, Alberta T2N 1N4, Canada. Electronic address: yeung@ucalgary.ca.
authors.forename: Edward C
authors.initials: EC
authors.lastname: Yeung
authors.affiliation: Department of Biological Sciences, 8888 University Dr., Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada. Electronic address: palma@neb.com.
authors.forename: Kristoffer
authors.initials: K
authors.lastname: Palma
authors.affiliation: Department of Biological Sciences, 8888 University Dr., Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada. Electronic address: kermode@sfu.ca.
authors.forename: Allison R
authors.initials: AR
authors.lastname: Kermode
mesh_terms: D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000378:metabolism; D057487:Enzyme Therapy; D006801:Humans; D010650:Phenylalanine Ammonia-Lyase / Q000008:administration & dosage / Q000378:metabolism* / Q000627:therapeutic use; D010661:Phenylketonurias / Q000188:drug therapy; D036226:Seedlings / Q000201:enzymology* / Q000502:physiology; D003313:Zea mays / Q000201:enzymology* / Q000502:physiology
chemical_list: D003451:Cryoprotective Agents; D010650:Phenylalanine Ammonia-Lyase
keywords: Enzyme substitution therapy; Freezing preservation; Japanese Striped corn; Oral enzyme therapeutic; Oral therapeutic; Phenylalanine ammonia lyase (PAL); Phenylketonuria (PKU); Seedling roots; Simulated digestion