Fingerprinting of Antenna Proteins of Photosystem I by Reversed Phase High Performance Liquid Chromatography

The paper reports the results of a study that has been performed to investigate the suitability of reversed phase high performance liquid chromatography (RP-HPLC) at producing confident fingerprints for rapid and simple identification of the light-harvesting proteins (Lhca) comprising the photosystem I (PSI) complex. The paper shows that Lhca proteins are completely resolved and separated from the other proteins comprising the core-complex by subjecting to RP-HPLC the entire PSI preparation. In each of all examined species the four antenna proteins elute in the same order, namely Lhca1 < Lhca3 < Lhca4 < Lhca2, with significant differences in retention times among the four proteins, indicating that each PSI light harvesting protein exhibits a unique chromatographic pattern reflecting hydrophobicity and stoichiometry of the PSI proteins. A good correlation between experimental retention times in RP-HPLC of proteins and their predicted values based on the overall hydrophobicity and polypeptide chain length of the examined antenna proteins is reported. These findings and the observation that the amino acid sequence of Lhca proteins are highly conserved among species, as well as the chromatographic behaviour in RP-HPLC of the PSI proteins are equivalent in all investigated species, endorse the possibility of using RP-HPLC as a highly confident fingerprint supportive of unequivocal assignment of the proteins in preparations from different species without the necessity of protein identification by additional methods and useful for comparison within a single and among different species for future studies of the PSI proteins.Acknowledgements We are grateful to Dr. Maria Bianchetti and Dr. Corrado Ciambella for technical assistance.