Amylose glycogen phosphorylase

Enzymatic Polymerization of Amylose with Glycogen Phosphorylase... 

The fact that glycogen phosphorylase can be used to polymerize amylose was first demonstrated by Schaffner and Specht [110] in 1938 using yeast phosphorylase. Shortly after, the same behavior was also observed for other phosphorylases from yeast by Kiessling [111, 112], muscles by Cori et al. [113], pea seeds [114] and potatoes by Hanes [115], and preparations from liver by Ostern and Holmes [116], Cori et al. [117] and Ostern et al. [118]. These results opened up the field of enzymatic polymerizations of amylose using glucose-1-phosphate as monomer, and can be considered the first experiments ever to synthesize biological macromolecules in vitro. 

The strict primer dependence of the glycogen phosphorylases makes them ideal candidates for the synthesis of hybrid structures of amylose with non-natural materials... 

The strict primer dependence of the glycogen phosphorylases makes them ideal candidates for the synthesis of hybrid structures of amylose with non-natural materials (e.g., inorganic particles and surfaces, synthetic polymers). For this, a primer functionality (maltooligosaccharide) can be coupled to a synthetic structure and subsequently elongated by enzymatic polymerization resulting in amylose blocks. 

Numerous analogs of carbohydrate polymers (i.e., amylose, glycogen) have been prepared from modified monosaccharide 1-phosphates with phosphorylase (Fig. 13-11 shows the natural substrates) l159 162l. 

Enzymatic polymerization of amylose with glycogen phosphorylase... 

In vivo linear a-l,4-glucans are synthesized from ADP-glucose by the enzyme glycogen synthase [94-97]. The enzyme, as well as the monomer, are quite sensitive and therefore most researchers (at least in the field of polymer science) prefer to use phosphorylase for the synthesis of amylose. 

The glycogen phosphorolysis of phosphorylase can be reverted, which makes it possible to enzymatically polymerize amylose as well as hybrid structures with amylose as outlined in the following section. 

Fig. 33. Experimentally observed molecular-weight dependence of (S2)z for amylose chains grafted onto glycogen (filled circles) and partially debranched amylopectin (open circles). Grafting was achieved by potato phosphorylase (15 rays for glycogen and 12000 per amylopectin molecule), and by muscle phosphorylase (400 rays per glycogen molecule)142,143)...

Formation of glycogen chains by glycogen muscle phosphorylase and starch amylose by potato phosphorylase. 

Earlier studies on the properties of phosphorylases isolated from various sources have indicated that their subunits are similar in size with about 100,000 daltons.15-17 The reaction proceeds in a rapid equilibrium random Bi-Bi mechanism as has been shown by kinetic studies with rabbit skeletal muscle phosphorylases a18-20 and b,21,22 rabbit liver enzyme,23 potato tuber enzyme,24 and the enzyme from E. coli.25) In contrast, the substrate specificities for various glucans differ considerably depending on the enzyme sources. The rabbit muscle enzyme has high affinity for branched glucans such as glycogen and amylopectin but low affinity for amylose and maltodextrin.26,27 The potato tuber enzyme can act on amylose, amylopectin, and maltodextrin but only poorly on glycogen,28,29 while the E. coli enzyme shows high affinity for maltodextrin.10 ... 

Muscle phosphorylase synthesizes an amylose from a-n-glucopyranosyl phosphate, whilst impure heart and liver preparations give polysaccharides with the physicochemical properties of a glycogen (ref. 27). 

Methods of assay (m) = methylation (p) = periodate oxidation (e) = enzymic. B.V. = Blue Value (compare Refs. 3 and 4). Xmax = wavelength of maximum absorption of polysaccharide-iodine complex. Priming activity toward P-enzyme soluble starch = 1.0. Priming activity toward muscle phosphorylase glycogen = 100, corn amylopectin = 63. Molecular weight, 64,000 (compared with about 100,000 for the parent amylose). 

The exact way of amylopectin biosynthesis in plants is still not known today. In our current research we are using a tandem reaction of two enzymes to synthesize artificial amylopectin or rather (hyper)branched amylose in vitro. One enzyme is responsible for building the linear (amylose) part while the other enzyme introduces die branches, phosphorylase and glycogen branching enzyme respectively. 

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