• Mannose 6 Phosphate
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Mannose-6-phosphate isomerase (3H1Y) from Salmonella typhimurium is involved with glycolysis and gluconeogenesis.  Mannose-6-phosphate isomerase is a monomer which can be found in the cytoplasm of the cell.  Under the phosphoglucose isomerase (PGI) superfamily, mannose-6-phosphate isomerase (MPI) acts as an enzyme for monosaccharide conversion.  MPIs are divided into two groups, type I and type II.  The mannose-6-phosphate isomerase bound to a substrate and metal atom is classified as a type I MPI.  The binding of metal ions increases binding of the substrate to the active site. 

The isomerase catalyzes the interconversion of D-mannose-6-phosphate to D-fructose-6-phosphate, an aldose and a ketose. (Wu)  When fructose-6-phosphate (F6P) is converted to mannose-6-phosphate (M6P), gluconeogenesis occurs and glucose is generated; the resulting M6P can also act as a cellular identifier for transport and membrane identification.  When the reverse occurs, the isomerase prepares fructose-6-phosphate for glycolysis.  Along with the conversions, the ring is opened and closed throughout the reaction.  His99 and Asp270 may catalyze the ring opening step.  The isomerization occurs due to acid/base catalysis with proton transfer between the first C atoms of the substrate.  MPI first binds by coordination to zinc; it’s electrophilic nature results in the transfer of the carbonyl oxygen double bond.  As the enzyme, zinc stabilizes the intermediate through charge neutralization. The zinc coordinates with the ligand carbonyl and hydroxyl oxygens on C-1 and C-2 to form the transient enediol intermediate.  (Gracy)  MPI has a significant pH activity range from 6.5-9.0 with its most optimal activity at pH of 8.5.

            In terms of structure, mannose-6-phosphate consists of three main domains: two β domains and a single α domain; the active site of the enzyme is located in the central β domain. (Sagurthi)  The mannose-6-phosphate isomerase secondary structure consists of 28 β sheet strands and 16 α helices; of the 16 helices, four are 3/10 helices.  MPI is 29% helical composition (114 residues) and 29% beta sheet (116 residues) with the total length of the isomerase counted at 393 residues. (PDB)  The molecular weight of MPI is 42748.8 Da and the estimated isoelectric point is a pH of 5.49.  (Expasy, 1) There are three specific ligands in the structure of MPI.  These include an 1,2-ethanediol unit, a fructose-6-phosphate ligand, and a zinc ion. 

            His99, Lys132, His131 and Asp270 are the nearest amino acids in the active site and are likely the groups that interact with the substrate for the proton transfer.  It is also suggested that the ring-opening step is catalyzed by His99 and Asp270.  These amino acids in a residue chain of approximately 130 in length interact with the Zn²⁺ and undergo a conformational change upon substrate binding.  Thermal denaturation also demonstrates that the metal binding stabilizes the protein with a higher temperature resistance when MPI is bound with a cation: Zn­²⁺, Mn²⁺, Co²⁺, Mg²⁺, and Ni²⁺. (Sagurthi)

            Structurally there are three domains to MPI: the catalytic, carboxyl, and helical domains.  The helical domain consists of alpha helices solely.  The catalytic, in contrast, consists of almost only beta sheets.  The catalytic domain, where the binding of the metal ion occurs, has a mixture of sheets and helices. 

            A similar protein, phosphomannose isomerase (1PMI), has a similar function to MPI, the reversible isomerization of fructose-6-phosphate and mannose-6-phosphate, using the similar mechanism. (Cleasby)  The Z-score of PMI is 42.4 as the basic structure is shared; however, there is only one zinc ligand found on the phosphomannose isomerase enzyme.  (DALI, 1)  In similar primary structure, mannose-6-phosphate isomerases share almost the exact sequence amongst different forms of bacteria.  99% of the sequence is shared between the MPI of Salmonella typhimurium, Salmonella enterica, Shigella flexneri, and Escherichia coli. (BLAST)

            Mannose-6-phosphate isomerase is essential for bacterial growth as mannose is the sole source of carbon.  MPI has been connected to the production of exopolysaccharide alginate, which coats bacteria and protects them from antibiotics.  Removal of MPI has been shown to cause an accumulation of mannose-6-phosphate in mice, which lead to toxic death.  MPI is important in this way as a drug target.