Structural features of the Nostoc punctiforme debranching enzyme reveal the basis of its mechanism and substrate specificity.
The debranching enzyme Nostoc punctiforme debranching enzyme (NPDE) from the cyanobacterium Nostoc punctiforme (PCC73102) hydrolyzes the alpha-1,6 glycosidic linkages of malto-oligosaccharides. Despite its high homology to cyclodextrin/pullulan (CD/PUL)-hydrolyzing enzymes from glycosyl hydrolase 13 family (GH-13), NPDE exhibits a unique catalytic preference for longer malto-oligosaccharides (>G8), performing hydrolysis without the transgylcosylation or CD-hydrolyzing activities of other GH-13 enzymes. To investigate the molecular basis for the property of NPDE, we determined the structure of NPDE at 2.37-A resolution. NPDE lacks the typical N-terminal domain of other CD/PUL-hydrolyzing enzymes and forms an elongated dimer in a head-to-head configuration. The unique orientation of residues 25-55 in NPDE yields an extended substrate binding groove from the catalytic center to the dimeric interface. The substrate binding groove with a lengthy cavity beyond the -1 subsite exhibits a suitable architecture for binding longer malto-oligosaccharides (>G8). These structural results may provide a molecular basis for the substrate specificity and catalytic function of this cyanobacterial enzyme, distinguishing it from the classical neopullulanases and CD/PUL-hydrolyzing enzymes.
Dumbrepatil
Arti Baban
AB
Department of Food Science and Technology, Chungbuk National University, Chungju, 361-763, Korea.
Choi
Ji-Hye
JH
Park
Jong Tae
JT
Kim
Myo-Jeong
MJ
Kim
Tae Jip
TJ
Woo
Eui-Jeon
EJ
Park
Kwan Hwa
KH
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D046937:Nostoc / Q000201:enzymology*; D011487:Protein Conformation; D055503:Protein Multimerization; D016415:Sequence Alignment; D013379:Substrate Specificity
D001426:Bacterial Proteins