1. Supplementary Table 1. Purification of Chi-V from E. coli Step Protein Activity Specific activity Yield (mg) (mmol/min) (mmol/min/mg) (%) Crude extract 2982.0 159.4 0.05 100 SP-Sepharose 53.24 62.3 1.17 39.1 Sephacryl 28.76 34.4 1.20 21.6 S-100 HR

2. 66.4 39.2 26.1 14.3 22.8 12 (kDa) Supplementary Fig. 1. Supplementary Fig. 1. SDS–PAGE was done by the method of Laemmli. Lane 1, marker proteins and lane 2, purified NtChiV. The gel was stained with Coomassie brilliant blue R250.

3. A B C D pHTemperature ( o C) 02468101220304050607080 40 60 20 100 80 40 60 20 100 0 0 Relative activity (%) Supplementary Fig. 2. Supplementary Fig. 2. Effects of pH and temperature on activity and stability of NtChiV. A, The optimum pH of the chitinase was measured after incubation with glycolchitin in the buffer at various pHs at 37 o C for 15 min. B and D, The pH and thermal stabilities of the enzyme were measured from the residual activities after incubation in the buffer at various pHs at 37 o C for 6 h and in 50 mM sodium acetate buffer, pH 5.0, at various temperatures for 6 h, respectively. C, The optimum temperature was determined by measuring the activities in 50 mM sodium acetate buffer, pH 5.0, at various temperatures.

4. 0 1 2 3 4 5 6 7 8 020406080100 120 0 1 2 3 4 5 6 7 8 020406080100120140160 180 Concentration (mM) Reaction Time (min) (NAG) 6 (NAG) 2 (NAG) 3 (NAG) 4 (NAG) 2 6 0 1 2 3 4 5 020406080100 120 (NAG) 5 (NAG) 2 (NAG) 3 A B C Supplementary Fig. 3. Time-courses of the enzymatic hydrolysis of (NAG) n. A, substrate (NAG) 6 ; B, substrate (NAG) 5 ; C, substrate (NAG) 4. Enzyme concentration was 0.16  M. The enzymatic reaction was conducted in 20 mM sodium acetate buffer pH 5.0 at 40 o C. Determinations of the products were performed by a gel-filtration HPLC using a column of TSK-GEL G2000PW (Tosoh). Symbols: squares, (NAG) 2 ; triangles, (NAG) 3 ; diamonds, (NAG) 4 ; circles, (NAG) 6. Lines were obtained by roughly following the experimental data points.

5. 100 Mass (m/z) A B C (NAG) 6 (NAG) 3 (NAG) 4 (NAG) 2 (NAG) 5 (NAG) 6 (NAG) 4 (NAG) 3 (NAG) 2 (NAG) 7 (NAG) 8 100 0 0 Intencity (%) 0 2004008001000120060014001600 14001600 14001600 0 10 (NAG) 7 (NAG) 8 0 Supplementary Fig. 4. MALDI-TOF-MS analysis of the enzymatic products. A, standards, (NAG) 2 ~(NAG) 6 ; B, substrate (NAG) 6 ; C, enzymatic products from the substrate (NAG) 6. The reaction mixture, consisting of 0.16  M NtChiV and 5.0 mM (NAG) 6 in 20 mM sodium acetate buffer, pH 5.0, was incubated for 10 min at 40 o C. The reaction mixture was mixed with an equal volume of 2,5- dihydroxy benzoic acid (2,5-DHB) and the resultant solution was employed for mass measurements using a MALDI micro MX (Waters). (NAG) 2-6 oligosaccharides were used as standards for m/z.