Using signal peptide prediction with caution, a case study in Aspergillus niger xylanase

Abstract Signal peptide (SP) prediction is used, but not known correct or not. The Aspergillus niger GH10 xylanase (XynB) SP is predicted to be 1–19 (Met 1 –Ser 19 ) residue, differing from the Penicillium simplicissimum xylanase 1–25 residue SP. To determine the real SP, two types of XynB, XynΔ19 and XynΔ25, were constructed by respectively deleting the 19 (Met 1 –Ser 19 ) or 25 (Met 1 –Arg 25 ) residues. The XynΔ25 had 10 °C higher T opt and 21.6-times longer thermostability than the XynΔ19 (46 vs. 36 °C and 47.6 vs. 2.2 m). When the kinetics were assayed, the XynΔ25 had ∼2.1-times higher V max and higher binding-affinity for xylan than the XynΔ19 (53.7 vs. 25.3 μmol/ml/m and 2.43 vs. 2.96 mg/ml). Thus, the XynB real SP is the 1–25 and not predicted 1–19 residues. The extra six N-terminal residues (Glu 20 Pro 21 Ile 22 Glu 23 Pro 24 Arg 25 ) drastically interfered with the XynΔ19 thermal activity, thermostability, and catalytic efficiency.