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abstracts
Ribosome Inactivating Proteins (RIPs) are capable of inhibiting protein synthesis by catalytically hydrolyzing at specific purine residues from the sarcin / ricin loop of large ribosomal RNA. There are two types of RIPs: type 1 - RIPs (RIP-1) are single polypeptide chain proteins while type 2 - RIPs contain two polypeptide chains which are covalently linked by a disulphide bond. Studies have indicated that stacking interactions play a dominant role in the ligand binding to RIPs. However, the structural basis of these interactions with RIPs as well as the nature of stack pairing and associated molecular mechanisms are not clearly understood. In order to examine the significance of these stacking interactions and the role of various aromatic residues involved, we have carried out the structural analysis of the complexes of RIP-1 from Momordica balsamina (MbRIP-1) and crystallized it with six different sugar molecules, ribose, fucose, glucose, fructose, maltose and lactose. The crystals belong to hexagonal space group H3 with approximate cell dimension of a = b = 130 Å, c = 40.5 Å. The r.m.s. deviation for the Calpha atoms in the complexed structures was found to be in the range of 0.5 Å to 0.9 Å. The aromatic rings of sugars are seen to be involved in stacking with aromatic rings of Tyr 70 and Tyr 111. The side chain torsion angles chi1 and chi2 of Tyr 70 in the complexes with ribose, fucose, glucose, fructose, maltose and lactose were found to be 64° / 161°, 166° / 86°, 6° / 172°, 70° / 167°, 64° / 161° and -75° / 178° respectively. The sugars are held firmly in the binding site of RIP-1 with the help of stacking interactions and hydrogen bonds. The results presented here have revealed the significance of aromatic stack pairing and delineated the stabilizing role of tyrosine residue in these interactions.
abstracts
Lactoperoxidase is a member of mammalian heme peroxidase superfamily that consists of lactoperoxidase (LPO), myloperoxidase (MPO), thyroid peroxidase (TPO), eosinophil peroxidase (EPO). So far crystal structures of only two mammalian peroxidases LPO and MPO have been determined. The structures of LPO have been determined from the samples obtained from bovine, buffalo, goat and sheep. Their complexes have also been determined with inhibitors and substrate analogues. However, the structures of EPO and TPO have not been obtained so far. Due to significant sequence identity between LPO and TPO, the structures of these proteins are expected to be similar. Similarly their substrates will also have similarity as well as the substrate and enzyme interactions will be similar. Therefore, a complex of LPO was prepared with propylthiouracil (PTU), a compound used as a drug in thyroid ailments. The complex was crystallized using co-crystallization. The crystals of native LPO were also soaked in the solution containing PTU. The crystals belonged to monoclinic space group with cell dimensions a = 80.2 Å, b = 82.5, c = 95.0, β = 73.7° There were four molecules of LPO in the asymmetric unit. The structure determination of the complex revealed that PTU binds to the LPO at the distal heme site. It is held at this site through several hydrogen bonds and van der Waals contacts. The mode of binding and number of interactions suggest that a similar mode of binding may occur with TPO. Based on the information of interactions and the missing potential interactions, the modifications in the structure of PTU are suggested so that the improved design of the TPO inhibitor is obtained.
