Samenvatting
Lectins are a group of carbohydrate-binding proteins or glycoproteins that are widely distributed in nature. Due to their ability to bind carbohydrates, lectins are thus powerful tools for the isolation and analysis of glycoconjugates. The gene encoding the seed lectin of three Pterocarpus species (P. indicus, P. echinatus and P. officinalis) (family: Leguminosae, subfamily: Papilionoideae, tribe: Dalbergieae) was sequenced, starting from the total genomic DNA and using primers originally designed for P. angolensis seed lectin, whose sequence was previously determined (NCBI protein sequence database, accession numbers CADI9803-11).
The sequence results yielded a DNA fragment without introns coding for a mature protein and a leader sequence (the latter was only determined in the case of P. officinalis and P. indicus).
The number of amino acid (241-252) of the mature protein corresponds to the molecular size of the lectins isolated from the Pterocarpus seeds (27-28 kDa, as determined by SDS-PAGE). It seems that in the case of the Pterocarpus seed lectins, no proteolytic processing occurs and two monomers come together and associate to form a dimer.
Alignment of the putative amino acid sequences from the three Pterocapus species and P. angolensis seed lectins reveals a high percent of homology. This result also indicated that P. indicus, P .echinatus and P .officinalis lectins have conserved sugar binding sites: the four essential residues (Asp86 in loop A, Glyl06 in the omega loop, and Asn138 and an aromatic residue Phe132 in the metal binding loop) are fully conserved. Moreover the amino acid preceding Asp 86 is also Ala, as in most legume lectins. All four Pterocarpus lectins have a putative glycosylation site at position Asn118-ThrI19-Ser120. Based on the known 3D-structure of the P. angolensis seed lectins (determined by X-ray crystallography) a reliable model could be built for the P. indicus, P. echinatus and P. officinalis lectin using the Swiss-model programs (http://swissmodel.expasy.org/). The structure obtained in all cases is completely similar to the structure of the P. angolensis lectin.
In this model the four amino acids that are known to be involved in sugar-binding (Glyl06, Asp86, Asn138 and Phe132) are occupying exactly the same position in space.
The P. indicus lectin contains a cysteine at position 176, but this residue is not likely to influence the sugar-binding properties. On the other hand, Ala221 in P. indicus, P. echinatus and P. officinalis (a Phe in P. angolensis) and Ser104 in P. officinalis (a Gly in P. angolensis), being in the immediate vicinity of the binding site, might affect the sugar-binding. These substitutions may be supposed to contribute to the marked differences in strength of adsorption of both lectins to the affinity matrix used for their purification.
The sequence results yielded a DNA fragment without introns coding for a mature protein and a leader sequence (the latter was only determined in the case of P. officinalis and P. indicus).
The number of amino acid (241-252) of the mature protein corresponds to the molecular size of the lectins isolated from the Pterocarpus seeds (27-28 kDa, as determined by SDS-PAGE). It seems that in the case of the Pterocarpus seed lectins, no proteolytic processing occurs and two monomers come together and associate to form a dimer.
Alignment of the putative amino acid sequences from the three Pterocapus species and P. angolensis seed lectins reveals a high percent of homology. This result also indicated that P. indicus, P .echinatus and P .officinalis lectins have conserved sugar binding sites: the four essential residues (Asp86 in loop A, Glyl06 in the omega loop, and Asn138 and an aromatic residue Phe132 in the metal binding loop) are fully conserved. Moreover the amino acid preceding Asp 86 is also Ala, as in most legume lectins. All four Pterocarpus lectins have a putative glycosylation site at position Asn118-ThrI19-Ser120. Based on the known 3D-structure of the P. angolensis seed lectins (determined by X-ray crystallography) a reliable model could be built for the P. indicus, P. echinatus and P. officinalis lectin using the Swiss-model programs (http://swissmodel.expasy.org/). The structure obtained in all cases is completely similar to the structure of the P. angolensis lectin.
In this model the four amino acids that are known to be involved in sugar-binding (Glyl06, Asp86, Asn138 and Phe132) are occupying exactly the same position in space.
The P. indicus lectin contains a cysteine at position 176, but this residue is not likely to influence the sugar-binding properties. On the other hand, Ala221 in P. indicus, P. echinatus and P. officinalis (a Phe in P. angolensis) and Ser104 in P. officinalis (a Gly in P. angolensis), being in the immediate vicinity of the binding site, might affect the sugar-binding. These substitutions may be supposed to contribute to the marked differences in strength of adsorption of both lectins to the affinity matrix used for their purification.
Originele taal-2 | English |
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Titel | Abstracts of the 194th meeting of the Belgian Society of Biochemistry and Molecular Biology (electronic) |
Pagina's | 48-48 |
Aantal pagina's | 1 |
Status | Published - dec. 2006 |