Overexpression, purification and characterization of the acidic ribosomal P-proteins from Candida albicans.
In all eukaryotic cells, acidic ribosomal P-proteins form a lateral protuberance on the 60S ribosomal subunit-the so-called stalk-structure that plays an important role during protein synthesis. In this work, we report for the first time a full-length cloning of four genes encoding the P-proteins from Candida albicans, their expression in Escherichia coli, purification and characterization of the recombinant proteins. Considerable amino acid sequence similarity was found between the cloned proteins and other known fungal ribosomal P-proteins. On the basis of their phylogenetic relationship and amino acid similarity to their yeast counterparts, the C. albicans P-proteins were named P1A, P1B, P2A and P2B. Using three different approaches, namely: chemical cross-linking method, gel filtration and two-hybrid system, we analyzed mutual interactions among the C. albicans P-proteins. The obtained data showed all the four P-proteins able to form homo-oligomeric complexes. However, the ones found between P1B-P2A and P1A-P2B were dominant forms among the C. albicans P-proteins. Moreover, the strength of interactions between particular proteins was different in these two complexes; the strongest interactions were observed between P1B and P2A proteins, and a significantly weaker one between P1A and P2B proteins.
Abramczyk
Dariusz
D
Department of Molecular Biology, Maria Curie-Skłodowska University, Institute of Microbiology and Biotechnology, Akademicka Street 19, 20-033 Lublin, Poland.
Tchórzewski
Marek
M
Krokowski
Dawid
D
Boguszewska
Aleksandra
A
Grankowski
Nikodem
N
D001483:Base Sequence; D002176:Candida albicans / Q000737:chemistry / Q000235:genetics; D003001:Cloning, Molecular; D005800:Genes, Fungal / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D010802:Phylogeny; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D016415:Sequence Alignment; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
D011994:Recombinant Proteins; D012269:Ribosomal Proteins; D001616:beta-Galactosidase