Understanding the Rate‐Limiting Step of Glycogenolysis by Using QM/MM Calculations on Human Glycogen Phosphorylase

Abstract

Liver glycogen phosphorylase (GP) is a key enzyme for human health, as its increased activity is associated with type 2 diabetes. The GP catalytic mechanism has been explored by quantum mechanics/molecular mechanics (QM/MM) methods. Herein, we propose a mechanism that proceeds by three steps: 1) it begins with transfer of a hydrogen atom from the phosphate group of the pyridoxal 5'-phosphate (HPO -PLP) cofactor to the phosphate substrate; 2) the glycosidic linkage is then cleaved through protonation of the glycosidic oxygen atom by a hydroxy group of the inorganic phosphate group; and 3) an oxygen atom of the phosphate performs a nucleophilic attack on the anomeric carbon atom of glucose, concomitant with the return of a proton from phosphate to PO -PLP, which finally leads to formation of the glucose-1-phosphate product and recovers the initial state of the PLP cofactor. The glycosidic bond cleavage and nucleophilic attack from the phosphate group to the glycosyl molecule have the highest activation free energies. The structural properties of the hereby characterized transition states could be very useful in structure-based drug design studies against liver GP.

An In Silico and an In Vitro Inhibition Analysis of Glycogen Phosphorylase by Flavonoids, Styrylchromones, and Pyrazoles

“…Pyrazoles 5, 6, and 7 hold alkyl chains at N-1 of the pyrazole moiety, namely, (CH 2 ) 9 CH 3 at pyrazole 5 and (CH 2 ) 11 CH 3 at pyrazoles 6 and 7. In 5-styrylpyrazoles (10)(11)(12)(13)(14)(15), four compounds hold a 2-hydroxyphenyl at C-3 of the pyrazole moiety (10)(11)(12)(13) and two compounds hold a phenyl tosylate (phenyl 4-methylbenzenesulfonate) at C-3 of the pyrazole moiety (14,15). Additionally, two compounds (10,14) hold an unsubstituted styryl group and the others present chloro (11) or methoxy (12,13,15) substituents.…”

“…In 5-styrylpyrazoles (10)(11)(12)(13)(14)(15), four compounds hold a 2-hydroxyphenyl at C-3 of the pyrazole moiety (10)(11)(12)(13) and two compounds hold a phenyl tosylate (phenyl 4-methylbenzenesulfonate) at C-3 of the pyrazole moiety (14,15). Additionally, two compounds (10,14) hold an unsubstituted styryl group and the others present chloro (11) or methoxy (12,13,15) substituents. Furthermore, different substitutions were assessed at N-1 of the pyrazole moiety, including the phenyl (12) and tosyl groups (13)(14)(15).…”

“…Under physiologic conditions, this enzyme catalyzes the degradation of glycogen, at the α-1,4-glycosidic linkage, to produce glucose 1-phosphate monomers in the presence of inorganic phosphate [10,11]. GP exists as a dimer, with two equal subunits of about 97 kDa [12]. The enzyme can be found in two different states, since phosphorylation of serine-14 by phosphorylase kinase (PhK) converts the inactive dephosphorylated form, GPb, into an active phosphorylated form, GPa [10].…”

“…The inhibition of skeletal mGP could lead to serious adverse effects in muscle functions, such as muscle cramping, which is related to the accumulation of glycogen in the tissue. Thus, the design of selective and effective inhibitors towards lGP remains a crucial topic of research [12,14].…”

Anthocyanins as Antidiabetic Agents—In Vitro and In Silico Approaches of Preventive and Therapeutic Effects

“…GP is an allosteric enzyme that changes between active (GPa) and tense (GPb) conformation states. Recently, the GP catalytic mechanism has been assessed at the atomistic level, and the characterized transition state geometries could be now used in structure-based drug design studies [ 156 ].…”