[A comparative structure-function analysis and molecular mechanism of action of endonucleases from Serratia marcescens and Physarum polycephalum].
Structural and functional characteristics were compared for wild-type nuclease from Serratia marcescens, which belongs to the family of DNA/RNA nonspecific endonucleases, its mutational forms, and the nuclease I-PpoI from Physarum polycephalum, which is a representative of the Cys-His box-containing subgroup of the superfamily of extremely specific intron-encoded homing DNases. Despite the lack of sequence homology and the overall different topology of the Serratia marcescens and I-PpoI nucleases, their active sites have a remarkable structural similarity. Both of them have a unique magnesium atom in the active site, which is a part of the coordinatively bonded water-magnesium complex involved in their catalytic acts. In the enzyme-substrate complexes, the Mg2+ ion is chelated by an Asp residue, coordinates two oxygen atoms of DNA, and stabilizes the transition state of the phosphate anion and 3'-OH group of the leaving nucleotide. A new mechanism of the phosphodiester bond cleavage, which is common for the Serratia marcescens and I-PpoI nucleases and differs from the known functioning mechanism of the restriction and homing endonucleases, was proposed. It presumes a His residue as a general base for the activation of a non-cluster water molecule at the nucleophilic in line displacement of the 3'-leaving group. A strained metalloenzyme-substrate complex is formed during hydrolysis and relaxes to the initial state after the reaction. The English version of the paper.
Shliapnikov
S V
SV
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, ul. Vavilova 32, Moscow, 117984 Russia.
Lunin
V V
VV
Blagova
E V
EV
Abaturov
L V
LV
Perbandt
M
M
Betzel
C
C
Mikhaĭlov
A M
AM
D000595:Amino Acid Sequence; D000818:Animals; D018360:Crystallography, X-Ray; D004706:Endodeoxyribonucleases / Q000737:chemistry* / Q000378:metabolism*; D004722:Endoribonucleases / Q000737:chemistry* / Q000378:metabolism*; D008274:Magnesium; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016839:Physarum polycephalum / Q000201:enzymology*; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D012706:Serratia marcescens / Q000201:enzymology*; D013329:Structure-Activity Relationship
D004706:Endodeoxyribonucleases; D004722:Endoribonucleases; C064579:I-Ppo endonuclease; C021835:Serratia marcescens nuclease; D008274:Magnesium