Structural and functional analysis of a low-temperature-active alkaline esterase from South China Sea marine sediment microbial metagenomic library.
A low-temperature-active alkaline esterase, Est12, from a marine sediment metagenomic fosmid library was identified. Est12 prefers short- and middle-chain p-nitrophenol esters as substrate with optimum temperature and pH value of 50 °C and 9.0, respectively, and nearly 50 % of maximum activity retained at 5 °C. The hydrolysis activity of Est12 was stable at 40 °C. Ca(2+) especially activated the activity of Est12 to about 151 % of the control. DEPC and PMSF inhibited the activity of Est12 to 34 and 25 %, respectively. In addition, Est12 was more tolerable to methanol compared to other organic solvents tested. The crystal structure of Est12 at 1.39 Å resolution showed that the cap domain which is composed of an α-helix and a flexible region resulted in a relatively wide spectrum of substrate, with p-nitrophenol caproate as the preferred one. Furthermore, the flexible cap domain and the high percentage of Gly, Ser, and Met may play important roles in the adaptation of Est12 to low temperature.
CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China.|Beijing Key Laboratory of Microbial Drug Resistance and Resistome, Beijing, 100101, China.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, 310006, China.
Yongfei
Y
Hu
Marine Bioengineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
Yinghui
Y
Liu
CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China.|Beijing Key Laboratory of Microbial Drug Resistance and Resistome, Beijing, 100101, China.
Jing
J
Li
Marine Bioengineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
Yanbin
Y
Feng
CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China.|Beijing Key Laboratory of Microbial Drug Resistance and Resistome, Beijing, 100101, China.
Na
N
Lu
CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China. zhubaoli@im.ac.cn.|Beijing Key Laboratory of Microbial Drug Resistance and Resistome, Beijing, 100101, China. zhubaoli@im.ac.cn.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, 310006, China. zhubaoli@im.ac.cn.
Baoli
B
Zhu
Marine Bioengineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China. xuesong@dicp.ac.cn.
Song
S
Xue
D000595:Amino Acid Sequence; D002118:Calcium / Q000378:metabolism; D002681:China; D018360:Crystallography, X-Ray; D004950:Esterases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D019015:Geologic Sediments / Q000382:microbiology*; D006863:Hydrogen-Ion Concentration; D056186:Metagenomics; D000432:Methanol; D008958:Models, Molecular; D009596:Nitrophenols / Q000737:chemistry / Q000378:metabolism; D009792:Oceans and Seas; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D013696:Temperature
D009596:Nitrophenols; D004950:Esterases; D002118:Calcium; D000432:Methanol; C024836:4-nitrophenol
Crystal structure; Esterase; Fosmid library; Low-temperature active; Marine sediment