Naslov
Essential role of peptidyl-prolyl isomerase sll0408 in Synechocystis sp. PCC 6803 development

Autori
Fulgosi, Hrvoje ; Ester, Lea ; Ljubešić, Nikola

Izvornik
Periodicum biologorum (0031-5362) 11 (2002), 4; 413-419

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
chaperone; chloroplast; cyanobacteria; photosynthesis; protein folding; TLP40

Sažetak
Background and purpose: The molecular chaperones, peptidyl-prolyl isomerases (PPIases), catalyse isomerization around peptide bonds which neighbour proline residues. They participate in a wide variety of important cellular processes, although their true physiological functions are still a subject of intense research. We recently identified a unique chloroplast PPIase, TLP40, which is involved in the regulation of oxygenic photosynthesis. Here, we describe the reverse genetic analysis of a cyanobacterial gene which could be the prokaryotic ancestor of TLP40 isomerase. Materials and Methods: The deduced amino acid sequence of the open reading frame sll0408 was analysed in detail and compared with related proteins. For the knock-out construct, the nucleotide sequence within the locus sll0408 was amplified using the polymerase chain reaction. The gentamycin-resistance cassette was inserted at a single BamHI site within the reading frame, and the construct was ligated with the pBluescript vector. The resulting pS3G-1 vector was transformed into Synechocystis sp. PCC6803 cells. Segregated resistant colonies were grown autotrophically and their ultrastructure was analysed using electron microscopy. Photosynthesis in mutant cell lines was measured with a Clark-type electrode. Results: The nucleotide sequence of the locus sll0408 predicts a protein of 402 amino acid residues (calculated Mr 43909). At the C-terminus, sll0408 displays high sequence similarity with the PPIase, TLP40, of higher-plant chloroplasts. All five fingerprint sequence motifs characteristic for this type of enzymes are conserved between the two proteins. slr0408 most likely belongs to the family of parvulines since it lacks the tryptophan residue responsible for the sensitivity of the isomerase to cyclosporin A. In analogy with TLP40, the N-terminal sequence of sll0408contains repeated leucines or isoleucines at every seventh position, which is a characteristic of the leucine zipper consensus motif in coiled-coil proteins. Cells lacking functional sll0408 (Dsll0408 line) have thylakoid membranes with irregular and segmented morphology. Electron-dense structures can be seen in the periplasmic space, often associated with the plasma membrane. Dsll0408 cells are severely impaired in photosynthesis, with rates of oxygen evolution almost equaling respiration. In contrast to wild-type cells of the same age, more than 90% of the Dsll0408 cells exhibit ultrastructural characteristics of advanced senescence. Conclusion: Locus sll0408 encodes a peptidyl-prolyl isomerase-like protein which shares high sequence similarity with the TLP40 isomerase of higher plant chloroplasts, indicating their functional and evolutional relatedness. sll0408 may be part of supramolecular chaperone complexes and its function appears to be essential for the maintenance of photosynthetic activity and proper development of Synechocystis cells.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

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