An extracellular D-(؊)-3-hydroxybutyrate oligomer hydrolase was purified from a poly(3-hydroxybutyrate)-degrading bacterium, Pseudomonas sp. strain A1. The purified enzyme hydrolyzed the D-(؊)-3-hydroxybutyrate dimer and trimer at similar rates. The enzyme activity was inhibited by a low concentration of diisopropylfluorophosphate. The molecular weight of the hydrolase was estimated to be about 70,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A 10-kbp DNA fragment of A1 was detected by hybridization with the gene (2 kbp) of an extracellular poly(3-hydroxybutyrate) depolymerase from Alcaligenes faecalis. Subsequent subcloning showed that a SmaI-KpnI fragment (2.8 kbp) was responsible for expression of the hydrolase in Escherichia coli and an in vitro transcription-translation system. The expressed protein detected by immunostaining had the same molecular weight as the purified enzyme from A1. The protein band detected in the in vitro transcription-translation system had a molecular size of 72 kDa. The nucleotide sequence of the SmaI-KpnI fragment was determined, and one open reading frame (2,112 nucleotides) was found. It specifies a protein with a deduced molecular weight of 72,876 (704 amino acids). In this sequence, the consensus sequence of serine-dependent hydrolysis, G-X-S-X-G, did not exist.Poly-3-hydroxybutyrate (PHB) is a unique intracellular reserve of organic carbon and/or chemical energy found in a wide variety of bacteria (6, 9) and is regarded as a potential biodegradable thermoplastic not derived from petroleum (12). Recently similar bacterial polyesters have been found, and these polyesters including PHB were called "polyhydroxyalkanoates" (PHAs) (7). Extracellular PHB metabolism is important in the application of these bacterial polyesters as biodegradable plastics. Many microorganisms that are capable of degrading PHB exist in the environment, and they secrete PHB depolymerases to degrade this polymer. There have been many studies on the extracellular PHB depolymerases from several bacteria and fungi (2,3,4,11,15,22,26,29). Genes for some enzymes were also cloned and sequenced (10,19,21). Alcaligenes faecalis T1, a PHB-degrading bacterium, secretes a D-(Ϫ)-3-hydroxybutyric acid oligomer hydrolase (3HB-oligomer hydrolase) (EC 3. 1.1.22) (23) besides a PHB depolymerase. Although the physiological role of the 3HB-oligomer hydrolase is not clear, it may be important to study this hydrolase in relation to extracellular degradation of PHB. Similar enzymes were found within the bacterial cells (8,16,25). Pseudomonas sp. strain A1 was isolated from activated sludge as a PHB-degrading bacterium (22) and was found to have an extracellular 3HB-oligomer hydrolase besides a PHB depolymerase. In this communication, we describe the purification and characterization of the 3HB-oligomer hydrolase and cloning and sequencing of its gene. MATERIALS AND METHODSBacterial strains, plasmids, and culture. Pseudomonas sp. strain A1 was isolated from activated sludge of the sewage treatment facility of...

ABSTRACT. The early response to the dehydration 15 (ERD15) gene is widely involved in the processes of signal transduction, programmed cell death, gene transcription, and stress tolerance in plants. In a previous study, the ERD15 gene was shown to be an important regulator of the abscisic acid response and salicylic acid-dependent defense pathway, acting as an important negative regulator of abscisic acid. The complete IbERD15 gene (accession No. KF723428) was isolated by reverse transcription-polymerase chain reaction. The IbERD15 gene contains an open reading frame of 504 bp, encodes a peptide of 167 amino acids, and has a molecular mass of 18.725 kDa. The transcript levels of the IbERD15 gene in a variety of tissues were examined by digital gene expression profiling. The roots of the sweet potato were treated by 3 degrees of polyethylene glycol, and the results indicate that the IbERD15 gene might play an important role in the defense response to drought stress. Moreover, the IbERD15 gene was successfully transformed into yeast cells for analysis of drought tolerance in transgenic yeast.