No slipping up: Banana proteome unpeeled using peptide libraries
- Published: Feb 1, 2013
- Author: Steve Down
- Channels: Proteomics & Genomics / Proteomics
Banana is an important fruit in the global economy for human nutrition, with total exports in the region of 17 million tonnes. This market is on the increase, due partly to the perception that it is a health food. The antioxidant and cell anti-proliferative activities have suggested that it might be able to protect against cardiovascular disease and cancer.
There have been a number of studies on the banana to identify metabolites in the fruit but there has been very little proteome activity. A knowledge of the protein composition of the Musa species could help to improve the quality of the fruit by developing new breeds as well as reveal the identities of biologically active peptides. The Global Musa Genomics Consortium has been working on the banana with the ultimate aim of improving its breeding and management and was involved in the recently announced sequencing of the banana genome.
One obstacle to proteomics studies of the fruit is the difficulty in extracting the proteins in the first place. Carbohydrates make up about 20% of fresh banana pulp whereas proteins constitute a lowly 1%. So, the medium-to-low abundant proteins, some of which might be important, can easily be left behind during conventional protein extraction procedures.
A group of European scientists has tackled this problem by employing combinatorial peptide ligand libraries. Pier Giorgio Righetti from the Miles Gloriosus Academy, Milan, with colleagues from Milan Polytechnic and the University of Alcalá, Madrid, devised a fairly complicated extraction protocol but were rewarded with a flood of proteins.
The banana pulp was initially extracted with either denaturing or non-denaturing buffer and each of these extracts was filtered before the proteins were precipitated out in bulk. Then a commercial peptide library comprising six hexapeptides was used to capture the proteins from each of the extracts at pH 2.2 and 7.4, representing physiological pH and mimicking reversed-phase conditions, respectively.
Following desorption from the beads, the proteins were separated by SDS-PAGE for analysis by HPLC-tandem mass spectrometry in the conventional manner. The identified peptides were searched against an expressed sequence tag database which contained data from three different species of the Musa family: Musa acuminata, Musa balbisiana and the hybrid of the two, known as Musa x paradisiacal. As a control, the precipitated proteins from the denaturing and non-denaturing steps were also dissolved in buffer and treated by SDS-PAGE.
More than 1000 proteins
The combined total of proteins from the three routes was 1131, with 679 identified uniquely from the ligand libraries and 282 from the controls. Only 170 proteins were common to the library and control groups, emphasising the ability of the ligand library to increase the amount of proteins that can be isolated.
The identified species included four of the five known banana allergens: profilin, class I chitinase, thaumatin-like protein and β-1,3-glucanase. Two other suspected allergens, a superoxide dismutase and a pectinesterase were also detected.
Of the other proteins, Righetti highlighted some that were involved in ripening processes. For instance, α-glucan water dikinase promotes starch degradation by assisting the transfer of phosphate into starch-like glucan. This process implies that the degradation of starch is linked to its phosphate content.
Also identified were proteins with antimicrobial and antibacterial properties, chaperones, protease inhibitors, folate receptors and small molecule transporters. The complete list of 1131 proteins was given in the supporting information in their report in Electrophoresis.
This is the first in-depth look at the banana fruit proteome and complements the work of the group on the proteomes of olive fruit and avocado fruit, more of the so-called recalcitrant plant systems from which it is difficult to extract proteins. Their efforts have illustrated the complexity of the banana proteome and will provide a basis for the production of special strains that might be resistant to disease and pathogens, to increase yields of this vital food crop, as well as strains with higher nutritional value.
Electrophoresis 2013, 34, 207-214: "In-depth proteomic analysis of banana (Musa spp.) fruit with combinatorial peptide ligand libraries"
Article by Steve Down
The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.