A mutant of phosphomannomutase1 retains full enzymatic activity, but is not activated by IMP: Possible implications for the disease PMM2-CDG

Abstract

The most frequent disorder of glycosylation, PMM2-CDG, is caused by a deficiency of phosphomannomutase activity. In humans two paralogous enzymes exist, both of them require mannose 1,6-bis-phosphate or glucose 1,6-bis-phosphate as activators, but only phospho-mannomutase1 hydrolyzes bis-phosphate hexoses. Mutations in the gene encoding phosphomannomutase2 are responsible for PMM2-CDG. Although not directly causative of the disease, the role of the paralogous enzyme in the disease should be clarified. Phosphomannomutase1 could have a beneficial effect, contributing to mannose 6-phosphate isomerization, or a detrimental effect, hydrolyzing the bis-phosphate hexose activator. A pivotal role in regulating mannose-1phosphate production and ultimately protein glycosylation might be played by inosine monophosphate that enhances the phosphatase activity of phosphomannomutase1. In this paper we analyzed human phosphomannomutases by conventional enzymatic assays as well as by novel techniques such as 31P-NMR and thermal shift assay. We characterized a triple mutant of phospomannomutase1 that retains mutase and phosphatase activity, but is unable to bind inosine monophosphate.

β-Glucose-1,6-Bisphosphate Stabilizes Pathological Phophomannomutase2 Mutants In Vitro and Represents a Lead Compound to Develop Pharmacological Chaperones for the Most Common Disorder of Glycosylation, PMM2-CDG

“…αG16P is degraded rapidly by PMM1 in particular in the presence of IMP [2,3]. We tested the stability of βG16P to the hydrolysis by PMM1 both in the absence and in the presence of IMP using 31 P-NMR.…”

“…αG16P itself could be considered an effective chaperone to rescue unstable pathological variants because it binds PMM2, induces the closure of the enzyme, raises the melting temperature, and activates catalysis. Regrettably, αG16P is hydrolyzed by PMM1 in particular in the presence of IMP [2,3]. Although data concerning the half-life of αG16P in vivo are not available, it can be predicted that its hydrolysis by PMM1 can hinder its utility as a drug.…”

“…Both enzymes require a sugar bisphosphate, α-mannose-1,6-bisphosphate or α-glucose-1,6-bisphospate, for their mutase activity. Besides being a mutase, PMM1 is phosphatase and plays a prevalent role in the degradation of esose bisphosphate in particular in the presence of inosine monophosphate (IMP) [2,3].…”

“…α-D-Glucose 1,6-bisphosphate (αG16P) could be considered an effective chaperone to rescue unstable pathological variants because it binds PMM2, induces the closure of the enzyme, raises the melting temperature, and activates catalysis [10,18,22]. However, it is rapidly hydrolyzed by PMM1 in particular in the presence of IMP [2,3].…”

CDG Therapies: From Bench to Bedside

“…PMM2-CDG (MIM: 212065) results from mutations in the gene encoding phosphomannomutase 2 (PMM2, EC 5.4.2.8), an enzyme that converts Man-6-P to mannose-1-phosphate (Man-l-P) in the cytoplasm. Phosphomannomutase 1 (PMM1) is a paralogous enzyme whose role, if any, in PMM2-CDG pathology has not been elucidated yet [ 191 , 192 , 193 ]. Prior to the identification of the underlying molecular cause of PMM2-CDG, Panneerselvam and Freeze had shown that the hypoglycosylation phenotype in PMM2-CDG patient fibroblasts could be resolved by the addition of Man to the culture medium [ 46 ].…”