**Qian Li, Fu Hu, Benwei Zhu \*, Yun Sun and Zhong Yao**

College of Food Science and Light Industry, Nanjing Tech University, Nanjing 211816, China; njlq@njtech.edu.cn (Q.L.); hufu@njtech.edu.cn (F.H.); sunyun\_food@njtech.edu.cn (Y.S.); yaozhong@njtech.edu.cn (Z.Y.)

**\*** Correspondence: zhubenwei@njtech.edu.cn; Tel.: +86-25-5813-9419

Received: 7 May 2019; Accepted: 24 May 2019; Published: 31 May 2019

**Abstract:** Alginate lyases have been widely used to prepare alginate oligosaccharides in food, agricultural, and medical industries. Therefore, discovering and characterizing novel alginate lyases with excellent properties has drawn increasing attention. Herein, a novel alginate lyase FsAlyPL6 of Polysaccharide Lyase (PL) 6 family is identified and biochemically characterized from *Flammeovirga* sp. NJ-04. It shows highest activity at 45 ◦C and could retain 50% of activity after being incubated at 45 ◦C for 1 h. The Thin-Layer Chromatography (TLC) and Electrospray Ionization Mass Spectrometry (ESI-MS) analysis indicates that FsAlyPL6 endolytically degrades alginate polysaccharide into oligosaccharides ranging from monosaccharides to pentasaccharides. In addition, the action pattern of the enzyme is also elucidated and the result suggests that FsAlyPL6 could recognize tetrasaccharide as the minimal substrate and cleave the glycosidic bonds between the subsites of −1 and +3. The research provides extended insights into the substrate recognition and degradation pattern of PL6 alginate lyases, which may further expand the application of alginate lyases.

**Keywords:** alginate lyase; polysaccharide lyase of family 6; characterization; degradation pattern
