Literature summary for 2.4.1.142 extracted from

Xu, X.X.; Li, S.T.; Wang, N.; Kitajima, T.; Yoko-O, T.; Fujita, M.; Nakanishi, H.; Gao, X.D.
Structural and functional analysis of Alg1 beta-1,4 mannosyltransferase reveals the physiological importance of its membrane topology (2018), Glycobiology, 28, 741-753 .

Search for more literature for 2.4.1.142

Cloned(Commentary)

Cloned (Comment)	Organism
gene Alg1, recombinant expression of GFP-tagged enzyme in Saccharomyces cerevisiae, recombinant expression of His6- or FLAG-tagged Alg1 mutants in Saccharomyces cerevisiae strain XGY29, in which chromosomal ALG1 is under the control of the glucose-repressible GAL1/10 promoter	Saccharomyces cerevisiae

Protein Variants

Protein Variants	Comment	Organism
K35A/K38A/K39A/R40A/V106I/M109L/V110I/V113I/V117I/I120L	site-directed mutagenesis, hydrophobic mut6 substitution of truncation mutant Alg1DELTA32 with additional mutations K35A, K38A, K39A, R40A. The mutant shows slower growth and decreased protein amount in membrane fraction compared to mutant Alg1NDELTA32-mut6A	Saccharomyces cerevisiae
additional information	deletion of the first 32 (alg1NDELTA32), 40 (alg1NDELTA40) or 50 (alg1NDELTA50) amino acids of Alg1p. Alg1NDELTA32 protein lacks the predicted N-terminal transmembrane domain, and further deletion of additional eight amino acids, including four positively charged residues, results in Alg1NDELTA40. Moreover, in Alg1NDELTA50, the first beta-sheet of Alg1 is completely eliminated. Mutant alg1NDELTA50 fails to rescue cell growth at the 30°C. Cells expressing alg1NDELTA32 show normal growth, similar to those expressing full-length Alg1, at normal temperature of 30°C and also relatively similar protein expression level. However, when the temperature is raised to 37°C, alg1NDELTA32-expressing cells show an obvious growth defect. The alg1NDELTA40-expressing cells exhibit a similar growth state as wild-type Alg1-expressing cells at 30°C, but partially affected growth at 37°C. Alg1NDELTA40 associates with the endoplasmic reticulum membrane in a nonperipheral manner, as does wild-type Alg1	Saccharomyces cerevisiae
S1A	site-directed mutagenesis, the N-terminal S2A-inserted Alg1p (FLAG-S2A-Alg1) is N-glycosylated by confirming the shift of protein band after PNGase treatment, whereas the C-terminal inserted form (Alg1-S2A-FLAG) is not	Saccharomyces cerevisiae
S1A/P20L	site-directed mutagenesis, no difference in N-glycosylation pattern after the replacement of proline residue is observed compared to the S2A mutant	Saccharomyces cerevisiae
V106I/M109L/V110I/V113I/V117I/I120L	site-directed mutagenesis, hydrophobic mut6 substitution of truncation mutant Alg1DELTA40, Alg1NDELTA40-mut6A is barely detected in either the membrane fraction or the cytosolic fraction, indicating that this mutant protein has been degraded. Alg1NDELTA40-mut6A mutant protein shows activity in vitro. Alg1NDELTA32-mut6A appears to function similar to intact Alg1 at 30°C and a similar expression level as wild-type Alg1	Saccharomyces cerevisiae

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
endoplasmic reticulum	ER, the predicted transmembrane domain localizes the N-terminus of Alg1 into the ER lumen. The N-terminal transmembrane domain including the following positively charged amino acids and an N-terminal amphiphilic-like alpha-helix domain exposed on the protein surface strictly coordinate the Alg1 orientation on the ER membrane. The N- and C-termini of Alg1 are located to luminal and cytosolic side of the ER, respectively. The conserved proline 20 residue has no effect on its membrane-spanning property. The positively charged amino acids (K35, K38, K39 and R40) downstream of the N-terminal transmembrane region are important for ER membrane attachment	Saccharomyces cerevisiae	5783	-
membrane	membrane topology of Alg1, overview. A conserved alpha-helix domain, Alg1 Nalpha3 domain, contributes to the membrane association of Alg1 protein	Saccharomyces cerevisiae	16020	-

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
GDP-alpha-D-mannose + N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	Saccharomyces cerevisiae	-	GDP + beta-D-mannosyl-(1->4)-N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	-	?
GDP-alpha-D-mannose + N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	Saccharomyces cerevisiae ATCC 204508	-	GDP + beta-D-mannosyl-(1->4)-N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	-	?

Organism

Organism	UniProt	Comment	Textmining
Saccharomyces cerevisiae	P16661	-	-
Saccharomyces cerevisiae ATCC 204508	P16661	-	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
GDP-alpha-D-mannose + N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	-	Saccharomyces cerevisiae	GDP + beta-D-mannosyl-(1->4)-N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	-	?
GDP-alpha-D-mannose + N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	-	Saccharomyces cerevisiae ATCC 204508	GDP + beta-D-mannosyl-(1->4)-N-acetyl-beta-D-glucosaminyl-(1->4)-N-acetyl-alpha-D-glucosaminyl-diphosphodolichol	-	?
additional information	Alg1 protein transfers a second beta-linkage mannose to the GlcNAc2-PP-Phy (phytanyl-diphosphoryl-alpha-N, N'-diacetylchitobioside) substrate	Saccharomyces cerevisiae	?	-	-
additional information	Alg1 protein transfers a second beta-linkage mannose to the GlcNAc2-PP-Phy (phytanyl-diphosphoryl-alpha-N, N'-diacetylchitobioside) substrate	Saccharomyces cerevisiae ATCC 204508	?	-	-

Synonyms

Synonyms	Comment	Organism
ALG1	-	Saccharomyces cerevisiae
Alg1p	-	Saccharomyces cerevisiae
beta-1,4 mannosyltransferase	-	Saccharomyces cerevisiae

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Saccharomyces cerevisiae

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.2	-	assay at	Saccharomyces cerevisiae

General Information

General Information	Comment	Organism
additional information	Alg1, a beta-1,4 mannosyltransferase (MTase), physically interacts with Alg2 (EC 2.4.1.132) and Alg11 (EC 2.4.1.257) proteins to form the multienzyme complex which catalyzes the addition of all five mannose to generate the Man5GlcNAc2-PP-Dol intermediate. Alg1 plays a central role in the formation of the multi-MTase, the topological information of Alg1 including the molecular mechanism of membrane association is analyzed. Alg1 shares structure similarity with Alg13/14 complex, which has been defined as a membrane-associated GT-B GTase, structural similarities, overview	Saccharomyces cerevisiae

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Release 2023.1 (January 2023)