Purification and characterization of an extracellular poly(L-lactic acid) depolymerase from a soil isolate, Amycolatopsis sp. strain K104-1.
Poly(L-lactic acid) (PLA)-degrading Amycolatopsis sp. strains K104-1 and K104-2 were isolated by screening 300 soil samples for the ability to form clear zones on the PLA-emulsified mineral agar plates. Both of the strains assimilated >90% of emulsified 0.1% (wt/vol) PLA within 8 days under aerobic conditions. A novel PLA depolymerase with a molecular weight of 24,000 was purified to homogeneity from the culture supernatant of strain K104-1. The purified enzyme degraded high-molecular-weight PLA in emulsion and in solid film, ultimately forming lactic acid. The optimum pH for the enzyme activity was 9.5, and the optimum temperature was 55 to 60 degrees C. The PLA depolymerase also degraded casein and fibrin but did not hydrolyze collagen type I, triolein, tributyrin, poly(beta-hydroxybutyrate), or poly(epsilon-caprolactone). The PLA-degrading and caseinolytic activities of the enzyme were inhibited by diisopropyl fluorophosphate and phenylmethylsulfonyl fluoride but were not significantly affected by soybean trypsin inhibitor, N-tosyl-L-lysyl chloromethyl ketone, N-tosyl-L-phenylalanyl chloromethyl ketone, and Streptomyces subtilisin inhibitor. Thus, Amycolatopsis sp. strain K104-1 excretes the unique PLA-degrading and fibrinolytic serine enzyme, utilizing extracellular polylactide as a sole carbon source.
Nakamura
K
K
Department of Molecular and Cell Biology, Graduate School of Agricultural Science, Tohoku University, Aoba-ku, Sendai 981-8555, Japan.
Tomita
T
T
Abe
N
N
Kamio
Y
Y
D000192:Actinomycetales / Q000201:enzymology* / Q000254:growth & development / Q000302:isolation & purification; D000595:Amino Acid Sequence; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D003470:Culture Media; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011091:Polyesters / Q000378:metabolism*; D012988:Soil Microbiology; D013379:Substrate Specificity; D013696:Temperature
D003470:Culture Media; D011091:Polyesters; C033616:poly(lactide); D002265:Carboxylic Ester Hydrolases; C425772:poly(lactic acid) depolymerase