Cofactor-independent phosphogycerate mutase

A glycerate-2,3-bisphosphate-independent phosphoglycerate mutase (PGAM), an enzyme typically found in plants, but also in some protists and in many bacteria has recently also been found in T. brucei. Most eukaryotes, including all vertebrates, have an isofunctional enzyme catalysing the same reaction, but their enzyme requires glycerate-2,3-bisphosphate as a cofactor and the two isofunctional enzymes are otherwise unrelated. Chevalier et al. (2000) have reported that the cofactor-independent enzyme of trypanosomes is closely related to the homologous enzymes from the plant cytosol. The analysis carried out in 2000 was repeated with similar results.

Further details of the analysis

The T. brucei PGAM was compared with the SwissProt release 39 / TrEMBL release 17 database indexed at European Bioinformatics Institute (EBI, Hinxton UK), containing 671,000 protein sequences, using the NCBI BLASTP program and the BLOSUM 62 matrix (http://www2.ebi.ac.uk/blastall/) . Click here to inspect the BLASTP output file.

The best E values were obtained with itself (Q9NG18) and with the sequence of the Solanum tuberosum (Potato) (Q9XE59), immediately followed by the PGAM from the green alga Chlamydomonas reinhardtii (AAK52421). Both the potato and C. reinhardtii sequence had 57% identical residues with the T. brucei sequence.

All PGAM sequences from the BLASTP output were aligned using the "RunDBClustalW" option in the BLASTP output. The ClustalW alignment is availabe here for inspection. From this alignment incomplete sequences and duplicates were removed, resulting in 35 sequences. In this alignment clearly two groups can be distinguised: the plant cytosolic PGAMs (the first 12 sequences), to which also the T. brucei PGAM (Q9NG18) belongs, and the bacterial and chloroplast PGAMs. This alignment was used to calculate pairwise distances . The removal of gap regions from the alignment has increased the percentages of identity, relative to the BLASTP output. The T. brucei PGAM (Q9XE59) has the highest identity (63%) or smallest distance (37%) to the PGAM of C. reinhartii (AAK52421). T. brucei PGAM is between 37 - 42% distant from the plant and algal sequences and between 61 -74% distant from the bacterial sequences.

A bootstrapped neighbor-joining tree was created using the program ClustalX with exclusion of all positions with gaps and with correction for multiple substitutions. T. brucei PGAM was monophyletic with the chlorophyte C. reinhardtii (70% bootstrap support) and robustly monophyletic with the alga and plant cytosolic PGAMs (100% bootstrap support).

Although likelihood mapping, as implemented in PUZZLE version 4.0.1, indicated the presence in the dataset of a strong phylogenetic signal (only 2.6% star-like quartets while 93.5 % of the quartet trees were well-resolved), a maximum likelihood tree created by PUZZLE gave a highly star-like tree. However, there was strong support for monophyly of T. brucei PGAM with that of C. reinhardtii and the plant cytosolic PGAMs. Four-cluster likelihood mapping to test the monophyly of the T. brucei PGAM (group a) with the algal and plant PGAMs (group b) as opposed to the prokaryotic PGAMs (groups c and d) showed that 100 % of trees supported the monophyly of T. brucei with the alga and plants.

Conclusion

The cofactor-independent PGAM of trypanosomes is closely related to the homologous enzymes from the plant cytosol.