High-performance liquid chromatography coupled on-line with electrospray ionization mass spectrometry for the simultaneous separation and identification of the Synechocystis PCC 6803 phycobilisome proteins

Zolla, Lello; Bianchetti, Maria

doi:10.1016/s0021-9673(01)00532-5

Cited by 37 publications

(23 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In Synechocystis sp. PCC 6803, investigated here, AP and PC are the dominant pigments, whereas PE is not present (Riethmann et al 1988;Zolla and Bianchetti 2001). The phycobilisomes content was measured according to Götz et al (1999).…”

Section: Measurements Of the Phycobilisomesmentioning

confidence: 99%

Comparison of two different modes of UV-B irradiation on synthesis of some cellular substances in the cyanobacterium Synechocystis sp. PCC6803

Marxen¹,

Vanselow²,

Hintze³

et al. 2010

J Appl Phycol

View full text Add to dashboard Cite

Two different modes of UV-B irradiation of the cyanobacterium Synechocystis sp. PCC 6803 are compared: turbidostatic control and additional physiostatic control. Under turbidostatic control, the cells were exposed to different constant UV-B irradiances, whereas under physiostatic control, an electronic control loop modulated UV-B irradiance in such a way that photosynthetic efficiency PSII was kept constant at a fixed set point. The UV-B-induced stimulation of the synthesis of pigments, α-tocopherol, and the antioxidative potential of methanolic soluble components of Synechocystis showed significant differences depending on the mode of irradiation, even though the overall doses were equal. For example, compared to the initial values, the concentrations of myxoxanthophyll and zeaxanthin increased to 226-244% and 453% upon constant UV-B irradiation in turbidostatic processes, whereas maxima of 600% and 740% were reached in turbidostatic process with additional physiostatic control. The α-tocopherol concentration increased under constant UV-B irradiances, up to a maximum of 150%. Under physiological control, however, maximum increases of 390% over the initial values were measured. Furthermore, a reaction scheme is given explaining the higher yield under physiostatic control.

show abstract

Section: Measurements Of the Phycobilisomesmentioning

confidence: 99%

Comparison of two different modes of UV-B irradiation on synthesis of some cellular substances in the cyanobacterium Synechocystis sp. PCC6803

Marxen¹,

Vanselow²,

Hintze³

et al. 2010

J Appl Phycol

View full text Add to dashboard Cite

show abstract

“…However, the UV spectroscopic techniques give just a rough estimation of the purity and nature of proteins, while SDS-PAGE provides only approximated values of the molecular weights of the proteins under study. Moreover, frequently the four main biliproteins that can be found in cyanobacteria (i.e., allophycocyanin-a, APC-a; allophycocyanin-b, APC-b; c-phycocyanin-a, CPC-a; and c-phycocyanin-b, CPC-b) [31] are not separated by SDS-PAGE due to their similar amino acid sequences and their similar molecular mass [32]. This limitation was observed by Campanella et al [33] who separated APC and CPC from S. platensis with low-performance LC and HPLC using UV detection.…”

Section: Introductionmentioning

confidence: 99%

“…Electron microscopy and crystallographic studies have also provided important information on structure and structure-function relationship of these phycobiliproteins [31] while ESI-MS has been used to analyze a purified solution containing CPCa and CPC-b [7]. More recently, a method based on HPLC-MS with the complete resolution of the protein components of phycobilisome from cyanobacterium Synechocystis 6803 was presented, which required more than 40 min per run [32].…”

Section: Introductionmentioning

confidence: 99%

Characterization of proteins from Spirulina platensis microalga using capillary electrophoresis‐ion trap‐mass spectrometry and capillary electrophoresis‐time of flight‐mass spectrometry

Simó¹,

Herrero²,

Neusüß

et al. 2005

Electrophoresis

View full text Add to dashboard Cite

In this work, a new capillary electrophoresis-mass spectrometry (CE-MS) procedure is developed to analyze proteins in Spirulina platensis microalgae. It is demonstrated that a fine optimization of several separation parameters is essential in order to achieve suitable CE-MS analysis of these proteins in natural extracts from microalgae. Namely, optimization of the composition of the separation buffer, electrospray conditions, and washing routine between runs are required in order to obtain reliable and reproducible CE-MS analyses of the main proteins found in this microalga (namely, allophycocyanin-alpha chain, allophycocyanin-beta, c-phycocyanin-alpha, and c-phycocyanin-beta). The relative molecular mass of these biopolymers is determined using two different MS instruments coupled to CE, i.e., CE-ion trap-MS and CE-time of flight-MS (CE-TOF-MS). A comparison between the results obtained with both instruments is carried out. The high resolution of the TOF-MS enables the distinction of small modifications in proteins and, thus, a more accurate mass determination. Interestingly, molecular mass values obtained by both CE-MS procedures agree very well while these experimental values are only in partial agreement with those theoretically expected (i.e., genetically derived masses). Some protein modifications due to amino acids exchange induced by nucleotide codon mutations are proposed to explain this difference.

show abstract

“…Other techniques as electron microscopy and crystallographic studies have also provided important information on structure and structure-function relationship of these phycobiliproteins [32], while ESI-MS has been used to analyze a purified solution containing CPCa and CPC-b [5]. More recently, a method based on HPLC-MS with the complete resolution of the protein components of phycobilisome from cyanobacterium Synechocystis 6803 was presented, which required more than 40 min per run [34].…”

Section: Introductionmentioning

confidence: 99%

Capillary electrophoresis-mass spectrometry ofSpirulina platensis proteins obtained by pressurized liquid extraction

Herrero¹,

Simó²,

Ibáñez³

et al. 2005

Electrophoresis

View full text Add to dashboard Cite

In this work, the usefulness of CE-MS to monitor and optimize the pressurized liquid extraction (PLE) of proteins from Spirulina platensis microalga is demonstrated. Crude and purified PLE extracts from microalga were analyzed by CE-MS. It was observed that the use of purification protocols of phycobiliproteins (namely, ultrafiltration or precipitation-dialysis-freeze drying) resulted in better CE resolution and MS signals, demonstrating that sample matrix plays an important role in CE-MS of proteins in real samples. Ultrafiltration was found less laborious and much faster than precipitation-dialysis-freeze drying (1 vs. 48 h). Direct analysis of crude extracts was demonstrated to be also possible by CE-MS, providing less-quality information but enough to characterize PLE extracts in a much faster way. Therefore, the latter protocol was selected to monitor and optimize the extraction process of phycobiliproteins from S. platensis. To do that, different extraction conditions were tested, including time, temperature and pressure of extraction, nature of pressurized liquid, distribution of microalga inside the extraction cell, type of packing, etc. It is demonstrated that the combined use of PLE and CE-MS allows the attainment of extracts rich in phycobiliproteins in short extraction times (namely, yields of 20% can be obtained in less than 2 h under the optimum PLE process in an automatic way). To our knowledge, this work shows for the first time the usefulness of CE-MS for monitoring and optimizing a PLE process.

show abstract

High-performance liquid chromatography coupled on-line with electrospray ionization mass spectrometry for the simultaneous separation and identification of the Synechocystis PCC 6803 phycobilisome proteins

Cited by 37 publications

References 30 publications

Comparison of two different modes of UV-B irradiation on synthesis of some cellular substances in the cyanobacterium Synechocystis sp. PCC6803

Comparison of two different modes of UV-B irradiation on synthesis of some cellular substances in the cyanobacterium Synechocystis sp. PCC6803

Characterization of proteins from Spirulina platensis microalga using capillary electrophoresis‐ion trap‐mass spectrometry and capillary electrophoresis‐time of flight‐mass spectrometry

Capillary electrophoresis-mass spectrometry ofSpirulina platensis proteins obtained by pressurized liquid extraction

Contact Info

Product

Resources

About