Paenidase, a novel D-aspartyl endopeptidase from Paenibacillus sp. B38: purification and substrate specificity.
We discovered and characterized a novel type D-aspartyl endopeptidase (DAEP) produced extracellularly by Paenibacillus sp. B38. This bacterial DAEP of M(r) 34,798 (named paenidase) appeared to be converted into a smaller form of M(r) 34,169 by the proteolytic removal of 5 amino acid residues from the N-terminal. The intact and modified forms of the enzyme displayed essentially the same enzymatic properties. The enzyme specifically hydrolyzed succinyl-D-aspartic acid alpha-(p-nitroanilide) and succinyl-D-aspartic acid alpha-(4-methylcoumaryl-7-amide) to generate p-nitroaniline and 7-amino-4-methylcoumarin, and internally cleaved a synthetic peptide (D-A-E-F-R-H-[D-Asp]-G-S-Y) of the [D-Asp](7) amyloid beta (Abeta) protein between [D-Asp](7)-G(8). Either was totally inert to the normal Abeta peptide sequence containing L-Asp, instead of D-Asp at the 7th position. Thus, paenidase is the first DAEP from a microorganism that specifically recognizes an internal D-Asp residue to cleave [D-Asp]-X peptide bonds.
Takahashi
Saori
S
Akita Research Institute of Food and Brewing, 4-26 Sanuki, Arayamachi, Akita 010-1623, Japan. saori@arif.pref.akita.jp
Ogasawara
Hironobu
H
Hiwatashi
Kazuyuki
K
Hori
Kazuyuki
K
Hata
Keishi
K
Tachibana
Tadanori
T
Itoh
Yoshifumi
Y
Sugiyama
Toshihiro
T
D000813:Anilides / Q000737:chemistry; D000814:Aniline Compounds / Q000138:chemical synthesis; D001224:Aspartic Acid / Q000031:analogs & derivatives / Q000737:chemistry; D016282:Aspartic Acid Endopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification*; D001406:Bacillaceae / Q000201:enzymology*; D003374:Coumarins / Q000138:chemical synthesis / Q000737:chemistry; D004789:Enzyme Activation / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity; D013696:Temperature
D000813:Anilides; D000814:Aniline Compounds; D003374:Coumarins; D011480:Protease Inhibitors; C515813:succinyl-aspartic acid alpha-(4-methylcoumaroyl-7-amide); C515812:succinyl-aspartic acid alpha-(4-nitroanilide); C019498:4-nitroaniline; D001224:Aspartic Acid; D016282:Aspartic Acid Endopeptidases; C028743:7-amino-4-methylcoumarin