Congenital Disorders of Glycosylation (CDG) are autosomal recessive defects in the synthesis of N-linked oligosaccharide chains. CDG group I (CDG-I) defects are defined as those caused by mutations in genes encoding enzymes used for the synthesis and transfer of lipid linked oligosaccharide (LLO) to newly synthesized proteins in the lumen of the ER. The steps in this pathway and the genes more ..
Target
| Sequence: MPI protein [Homo sapiens] Protein Family: Alpha-Mannosidase Binding Domain of Atg19/34 Gene:MPI Related Protein 3D Structures |
Depositor Specified Assays
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| AID | Name | Type | Comment |
|---|---|---|---|
| 1020 | Counter Screen for Glucose-6-Phosphate Dehydrogenase-based Primary Assay | screening | Counter Screen for Glucose-6-Phosphate Dehydrogenase-based Primary Assay |
| 1209 | HTS identification of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay. | confirmatory | HTS identification of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay |
| 1217 | uHTS Identification of Diaphorase Inhibitors and Chemcical Oxidizers: Counter Screen for Diaphorase-based Primary Assays | screening | uHTS Identification of Diaphorase Inhibitors and Chemcical Oxidizers: Counter Screen for Diaphorase-based Primary Assays |
| 1220 | HTS identification of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay using a high concentration of mannose 6-phosphate | confirmatory | HTS identification of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay using a high concentration of mannose 6-phosphate |
| 1535 | Confirmation of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay. | confirmatory | Confirmation of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay. |
| 1536 | Confirmation of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay using a high concentration of mannose 6-phosphate. | confirmatory | Confirmation of compounds inhibiting phosphomannose isomerase (PMI) via a fluorescence intensity assay using a high concentration of mannose 6-phosphate. |
| 1553 | Screening for Phosphomannose Isomerase inhibitors in cellular based assay using Hela cells. | other | Screening for Phosphomannose Isomerase inhibitors in cellular based assay using Hela cells. |
| 1620 | Toxicity Screening of PMI Inhibitors in Hela cells | other | Toxicity Screening of PMI Inhibitors in Hela cells |
| 1655 | Counter screen SAR assay for PMM2 inhibitors via a fluorescence intensity assay | confirmatory | Counter screen SAR assay for PMM2 inhibitors via a fluorescence intensity assay |
| 1666 | SAR assay for compounds that inhibit PHOSPHO1 | confirmatory | SAR assay for compounds that inhibit PHOSPHO1 |
Description:
Data Source: Sanford-Burnham Center for Chemical Genomics (SBCCG)
Source Affiliation: Sanford-Burnham Medical Research Institute (SBMRI, San Diego, CA)
Network: NIH Molecular Libraries Screening Centers Network (MLSCN)
Grant Number: R03 MH082386-01
Assay Provider: Dr. Hudson H. Freeze, Sanford-Burnham Medical Research Institute, San Diego, CA
Congenital Disorders of Glycosylation (CDG) are autosomal recessive defects in the synthesis of N-linked oligosaccharide chains. CDG group I (CDG-I) defects are defined as those caused by mutations in genes encoding enzymes used for the synthesis and transfer of lipid linked oligosaccharide (LLO) to newly synthesized proteins in the lumen of the ER. The steps in this pathway and the genes encoding them are very similar from yeast to human. It requires 30-40 single gene products, each dependent on the previous step in the linear sequence to produce and transfer the LLO to protein. Therefore, mutations in any step may cause a type of CDG. There is considerable overlap in the clinical presentations between different types of CDG and a broad diversity within each type. The most common form of CDG, called Type Ia (CDG-Ia), is caused by defects in PMM2 (Man-6-P to Man-1-P), the gene that encodes phosphomannomutase. Mortality is 20% in the first 5 yrs, but then patients stabilize. Currently, there is no treatment for the CDG-Ia.
CDG-Ib patients, who are deficient in phosphomannose isomerase (PMI) catalyzing conversion of Man-6-P to Fru-6-P, are successfully treated with free mannose. Unfortunately, mannose therapy is not effective for CDG-Ia patients, most likely due to efficient Man-6-P consumption in the PMI reaction. It is believed that patients with Congenital Disorder of Glycosylation Type Ia (CDG-Ia) will benefit from dietary mannose if there is a simultaneous reduction of phosphomannose isomerase (PMI) activity. This would allow a modest intracellular accumulation of Man-6-P and drive metabolic flux into the glycosylation pathway using the residual PMM2 activity. It is assumed that a non-competitive inhibitor would work best in this setting; however, identification of chemical probes with diverse modes of action (MOA) would be advantageous for further characterization of PMI and PMM variants.
The purpose of this assay is to identify inhibitors of human PMI. This is accomplished by using a G6PD- NADPH-coupled assay. In the assay PMI activity is detected through conversion of its product, fructose-6-phosphate, to glucose-6-phosphate catalyzed by phosphoglucose isomerase (PGI) and subsequent oxidation of glucose-6-phosphate to 6-phosphogluconolactone concomitant with NADP-to-NADPH conversion catalyzed by glucose-6-phosphate dehydrogenase (G6PDH). The NADPH is then detected via a resazurin-diaphorase fluorogenic reaction.
AIDs 1020, 1209, 1217, 1220, 1535, 1536, 1553, 1620, 1655 and 1666 were carried out to select probe molecules SID57309177(ML096) and SID57287553(ML089).
Source Affiliation: Sanford-Burnham Medical Research Institute (SBMRI, San Diego, CA)
Network: NIH Molecular Libraries Screening Centers Network (MLSCN)
Grant Number: R03 MH082386-01
Assay Provider: Dr. Hudson H. Freeze, Sanford-Burnham Medical Research Institute, San Diego, CA
Congenital Disorders of Glycosylation (CDG) are autosomal recessive defects in the synthesis of N-linked oligosaccharide chains. CDG group I (CDG-I) defects are defined as those caused by mutations in genes encoding enzymes used for the synthesis and transfer of lipid linked oligosaccharide (LLO) to newly synthesized proteins in the lumen of the ER. The steps in this pathway and the genes encoding them are very similar from yeast to human. It requires 30-40 single gene products, each dependent on the previous step in the linear sequence to produce and transfer the LLO to protein. Therefore, mutations in any step may cause a type of CDG. There is considerable overlap in the clinical presentations between different types of CDG and a broad diversity within each type. The most common form of CDG, called Type Ia (CDG-Ia), is caused by defects in PMM2 (Man-6-P to Man-1-P), the gene that encodes phosphomannomutase. Mortality is 20% in the first 5 yrs, but then patients stabilize. Currently, there is no treatment for the CDG-Ia.
CDG-Ib patients, who are deficient in phosphomannose isomerase (PMI) catalyzing conversion of Man-6-P to Fru-6-P, are successfully treated with free mannose. Unfortunately, mannose therapy is not effective for CDG-Ia patients, most likely due to efficient Man-6-P consumption in the PMI reaction. It is believed that patients with Congenital Disorder of Glycosylation Type Ia (CDG-Ia) will benefit from dietary mannose if there is a simultaneous reduction of phosphomannose isomerase (PMI) activity. This would allow a modest intracellular accumulation of Man-6-P and drive metabolic flux into the glycosylation pathway using the residual PMM2 activity. It is assumed that a non-competitive inhibitor would work best in this setting; however, identification of chemical probes with diverse modes of action (MOA) would be advantageous for further characterization of PMI and PMM variants.
The purpose of this assay is to identify inhibitors of human PMI. This is accomplished by using a G6PD- NADPH-coupled assay. In the assay PMI activity is detected through conversion of its product, fructose-6-phosphate, to glucose-6-phosphate catalyzed by phosphoglucose isomerase (PGI) and subsequent oxidation of glucose-6-phosphate to 6-phosphogluconolactone concomitant with NADP-to-NADPH conversion catalyzed by glucose-6-phosphate dehydrogenase (G6PDH). The NADPH is then detected via a resazurin-diaphorase fluorogenic reaction.
AIDs 1020, 1209, 1217, 1220, 1535, 1536, 1553, 1620, 1655 and 1666 were carried out to select probe molecules SID57309177(ML096) and SID57287553(ML089).
Protocol
Please see pertinent AIDs: 1020, 1209, 1217, 1220, 1535, 1536, 1553, 1620, 1655, 1666.
Comment
Probe molecules are defined as the positives of this assay and assigned a score of 100.
Result Definitions
| TID | Name | Description | Histogram | Type | Unit | |
|---|---|---|---|---|---|---|
| Outcome | The BioAssay activity outcome | Outcome | ||||
| Score | The BioAssay activity ranking score |  | Integer | |||
| 1 | ML# | ML# of the compound | String |
Additional Information
Grant Number: R03 MH082386-01
Data Table (Concise)
Classification
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