Characterisation of general proteolytic, milk clotting and antifungal activity of Ficus carica latex during fruit ripening.

BACKGROUND: The physiological role of fig latex is to protect the plant from pathogens. Latex is a rich source of proteases, predominantly ficin. Fig latex also contains collagenolytic protease and chitinolytic enzymes. Our aim was to investigate changes in protein composition, enzyme and antifungal activities of fig latex during fruit ripening. RESULTS: Comparison of latex samples in different time periods showed a uniform increase of protein concentration in chronological order. The content of collagenolytic protease did not differ significantly in the latex samples, while the content of ficin decreased. Ficin-specific activity towards casein was the highest at the beginning of fruit development (about 80 U mg(-1)). Specific milk clotting activity increased as well as the abundance of casein band in the clots. Specific chitinolytic activity at the beginning of flowering was 6.5 times higher than the activity in the period when fruits are ripe. Antifungal activity is the most extensive in spring. CONCLUSION: Ficin forms with different casein specificities are present in different proportions during fruit ripening, which is of importance for applications in the dairy industry. The protection mechanism against insects and fungi, which relies on chitinolytic activity, is the most important in the early phases of flowering and is replaced with other strategies over time.

Identification, purification and characterization of a novel collagenolytic serine protease from fig (Ficus carica var. Brown Turkey) latex.

A novel collagenolytic serine protease was identified and then purified (along with ficin) to apparent homogeneity from the latex of fig (Ficus carica, var. Brown Turkey) by two step chromatographic procedure using gel and covalent chromatography. The enzyme is a monomeric protein of molecular mass of 41 ± 9 kDa as estimated by analytical gel filtration chromatography. It is an acidic protein with a pI value of approximately 5 and optimal activity at pH 8.0-8.5 and temperature 60°C. The enzymatic activity was strongly inhibited by PMSF and Pefabloc SC, indicating that the enzyme is a serine protease. The enzyme showed specificity towards gelatin and collagen (215 GDU/mg and 24.8 CDU/mg, respectively) and non-specific protease activity (0.18 U/mg against casein). The enzyme was stable and retained full activity over a broad range of pH and temperature. The fig latex collagenolytic protease is potentially useful as a non-microbial enzyme with collagenolytic activity for various applications in the fields of biochemistry, biotechnology and medicine.

Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein.

OBJECTIVE: Human BPIFA2 (parotid secretory protein) is a ubiquitous soluble salivary protein, which belongs to the PLUNC family of proteins. Having sequence similarity to bactericidal/permeability-increasing protein and lipopolysaccharide-binding protein, PLUNC proteins are probably involved in local antibacterial response at mucosal sites, such as oral cavity. The aim of the study was to isolate and characterize human BPIFA2. DESIGN: In this paper, we report one-step affinity chromatography method for BPIFA2 purification from whole human saliva. The isolated BPIFA2 was identified by trypsin mass fingerprinting and characterized by electrophoretic methods. Antibacterial activity of BPIFA2 against model microorganism Pseudomonas aeruginosa was shown in minimum inhibitory concentration and time kill study assays. RESULTS: The protein showed microheterogeneity, both in molecular weight and pI value. BPIFA2 inhibited the growth of P. aeruginosa in microgram concentration range determined by minimum inhibitory concentration assay. In the time kill study, 32µg/mL BPIFA2 showed clear bactericidal activity and did not cause any aggregation of bacteria. CONCLUSION: Affinity chromatography is well suited for isolation of functional BPIFA2 with a potent bactericidal activity against P. aeruginosa.