Selecting an appropriate method for expressing S locus F-box-S2 recombinant protein

- Understanding the molecular basis of self-incompatibility (SI) is essential for commercial production of fruit crops such as Apple.
- To investigate the molecular interactions in self-incompatibility locus (Slocus), the knowledge of tertiary structures of both male (i.e. S-locus F-box) and female (i.e. SRNase) proteins are necessary.
- The tertiary structure of male determinant (S locus F-box, SLF/SFB) remains unresolved, which could mainly be due to difficulties associated with its expression in the recombinant expression systems.
- This study demonstrates an approach for efficient expression of S locus F-box recombinant proteins for future functional and structural studies.

A single locus (S locus) including at least two linked genes (female and male determinants) genetically controls the gametophytic self-incompatibility (GSI) in apple, which has evolved to avoid self-fertilization. There has been extensive work done on the female determinant of self-incompatibility, which has led to the determination of the tertiary structure of S-RNase. However, the tertiary structure of male determinant (S locus F-box, SLF/SFB) remains unresolved, which could mainly be due to difficulties associated with its expression in the recombinant expression systems. In addressing this, we have evaluated several in vivo (prokaryotic and eukaryotic) and in vitro expression systems for their efficiency in the expression of apple SLF2. The most successful expression of SLF2 (1 mg/ml) was achieved in E. coli using the synthesized gene in a high salt culture and applying heat shock before induction of culture. We therefore present an approach for the efficient expression of S locus F-box recombinant proteins for future functional and structural studies.