Cloning and expression of an extracellular-agarase gene from Streptomyces coelicolor A3(2) in Streptomyces lividans 66

Kendall, Kevin; Cullum, John

doi:10.1016/0378-1119(84)90060-x

Cited by 97 publications

(46 citation statements)

References 11 publications

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“…The pH optimum for activity of rAgaA (0.02 U/ml) was examined at 50 C in 50 mM Britton-Robinson universal buffers (pH [3][4][5][6][7][8][9][10][11][12]. The enzyme showed an optimal pH of about 7.0.…”

Section: Effects Of Ph On Activity and Stabilitymentioning

confidence: 99%

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Enzymatic Properties and Nucleotide and Amino Acid Sequences of a Thermostable β-Agarase from the Novel Marine Isolate, JAMB-A94

Ohta¹,

Nogi²,

Miyazaki³

et al. 2004

Bioscience, Biotechnology, and Biochemistry

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Section: Effects Of Ph On Activity and Stabilitymentioning

confidence: 99%

“…2) Agar-degrading bacteria isolated from marine and other environments so far have been assigned to the genera Pseudomonas, 3) Pseudoalteromonas, 4) Streptomyces, 5) Alteromonas, 6) Microscilla, 7,8) Vibrio, 9,10) and Cytophaga.…”

mentioning

confidence: 99%

Enzymatic Properties and Nucleotide and Amino Acid Sequences of a Thermostable β-Agarase from the Novel Marine Isolate, JAMB-A94

Ohta¹,

Nogi²,

Miyazaki³

et al. 2004

Bioscience, Biotechnology, and Biochemistry

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“…Several genes encoding extracellular enzymes have been cloned from Streptomyces species. These include endoglycosidase H (25), tyrosinase (16), agarase (4,17), P-lactamase (8), a-amylase (2,14,20,21,32,33), and xylanase (15,22). We have cloned in Streptomyces lividans the gene encoding a-amylase of Streptomyces griseus IMRU 3570 (31), a strain which is known to overproduce several extracellular enzymes (7).…”

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confidence: 99%

Characterization, expression in Streptomyces lividans, and processing of the amylase of Streptomyces griseus IMRU 3570: two different amylases are derived from the same gene by an intracellular processing mechanism

et al. 1991

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Extracellular amylase in Streptomyces lividans was undetectable in starch-supplemented medium. However, S. lividans produced fivefold-higher levels of amylase than Streptomyces griseus IMRU 3570 when transformed with the S. griseus amy gene. Two major proteins of 57 and 50 kDa with amylase activity accumulated in the culture broths of the donor S. gniseus and S. lividans transformed with the amy gene. Both proteins were also present in protoplast lysates in the same relative proportion; they gave a positive reaction with antibodies against the 57-kDa amylase. They did not differ in substrate specificity or enzyme kinetics. The two amylases were purified to homogeneity by a two-step procedure. Both proteins showed the same amino-terminal sequence of amino acids, suggesting that both proteins are derived from the same gene. The deduced signal peptide has 28 amino acids with two positively charged arginines near the amino-terminal end. When an internal NcoI fragment was removed from the amy gene, the resulting S. lividans transformants did not synthesize any of the two amylase proteins and showed no reaction in immunoblotting. Formation of the 50-kDa protein was observed when pure 57-kDa amylase was treated with supernatants of protoplast lysates but not when it was treated with membrane preparations, indicating that the native 57-kDa amylase could be processed intracellularly.A large number of extracellular enzymes are produced by Streptomyces species that are used by these saprophytic microorganisms to degrade polymeric nutrients in soil originating from decaying plant material. Several genes encoding extracellular enzymes have been cloned from Streptomyces species. These include endoglycosidase H (25), tyrosinase (16), agarase (4, 17), P-lactamase (8), a-amylase (2,14,20,21,32,33), and xylanase (15,22). We have cloned in Streptomyces lividans the gene encoding a-amylase of Streptomyces griseus IMRU 3570 (31), a strain which is known to overproduce several extracellular enzymes (7). It contains an open reading frame of 1,698 nucleotides which encodes a protein with a deduced molecular mass of 59,713 Da.Very little is known, however, about the mechanisms of posttranslational modification and secretion of extracellular enzymes of Streptomyces species. Secretion of extracellular enzymes in Bacillus species and other bacteria proceeds by membrane anchoring of the amino-terminal end of the newly synthesized intracellular protein (24). The interaction of the amino-terminal hydrophobic region of the protein (the leader peptide) produces a modification of the protein phospholipid arrangement of the lipid bilayer that results in the opening of channels through which the protein is secreted. Passage of the extracellular protein through the secretion channels results in the cleavage of the leader peptide by the signal peptidase (23,24).In Streptomyces species, the leader peptides of extracellular proteins are poorly known, and no detailed studies on the protein-processing mechanisms prior to or during the secretion have bee...

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“…Genes encoding endoglycosidase H from Streptomyces plicatus (52,53), agarase from Streptomyces coelicolor (8,29), a-amylase from Streptomyces hygroscopicus (26,39), and xylanases from Streptomyces sp. strain 36a (27) and Streptomyces lividans (43) have been cloned, and the nucleotide sequences of the first three have been determined (see references 52,12, and 26, respectively).…”

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alpha-Amylase gene of Streptomyces limosus: nucleotide sequence, expression motifs, and amino acid sequence homology to mammalian and invertebrate alpha-amylases

Long¹,

Virolle²,

Chang³

et al. 1987

J Bacteriol

117

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The nucleotide sequence of the coding and regulatory regions of the ca-amylase gene (aml) of Streptomyces limosus was determined. High-resolution Si mapping was used to locate the 5' end of the transcript and demonstrated that the gene is transcribed from a unique promoter. The predicted amino acid sequence has considerable identity to mammalian and invertebrate ca-amylases, but not to those of plant, fungal, or eubacterial origin. Consistent with this is the susceptibility of the enzyme to an inhibitor of mammalian a-amylases. The amino-terminal sequence of the extracellular enzyme was determined, revealing the presence of a typical signal peptide preceding the mature form of the a-amylase.Streptomycetes are gram-positive mycelial soil bacteria that obtain nutrients from macromolecular organic debris by using a wide range of hydrolytic enzymes. These enzymes include proteases, nucleases, lipases, and a variety of enzymes that hydrolyze different types of often quite complex polysaccharides (71).The last class of secreted enzyme has received considerable recent attention. Genes encoding endoglycosidase H from Streptomyces plicatus (52, 53), agarase from Streptomyces coelicolor (8, 29), a-amylase from Streptomyces hygroscopicus (26, 39), and xylanases from Streptomyces sp. strain 36a (27) and Streptomyces lividans (43) have been cloned, and the nucleotide sequences of the first three have been determined (see references 52, 12, and 26, respectively). Some of these genes are convenient for the study of gene expression and its regulation in streptomycetes, and all have potential application for the construction of expression and secretion vectors for the production of heterologous proteins in this industrially familiar genus.We recently reported the cloning of an a-amylase gene (am!) equivalent regions in other bacterial genes, including those involved in polysaccharide hydrolysis. Determination of the amino-terminal sequence of the extracellular form of the a-amylase protein enabled us to deduce the signal peptide sequence involved in the secretion process. a-Amylases are widely distributed among both procaryotes and eucaryotes. Substantial nucleotide and amino acid sequence data on these genes and proteins are now available. A previous comparison of the amino acid sequences of 11 different at-amylases was made by Nakajima et al. (46). While considerable amino acid sequence identity could be detected between six mammalian a-amylases (ranging from 80 to 90%), minimal (<10%) identity was detected between these enzymes and ax-amylases from a wide range of other species, including several bacteria, Aspergillus oryzae, and barley. Furthermore, little similarity could be detected between the a-amylases of these additional genera. In a comparison that included a-amylases from a wider variety of genera, we observed considerable amino acid sequence identity between the a-amylase of S. limosus and those of mammalian and invertebrate origin.Previous a-amylase amino acid sequence comparisons (46, 54) recognized four conserv...

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Cloning and expression of an extracellular-agarase gene from Streptomyces coelicolor A3(2) in Streptomyces lividans 66

Cited by 97 publications

References 11 publications

Enzymatic Properties and Nucleotide and Amino Acid Sequences of a Thermostable β-Agarase from the Novel Marine Isolate, JAMB-A94

Enzymatic Properties and Nucleotide and Amino Acid Sequences of a Thermostable β-Agarase from the Novel Marine Isolate, JAMB-A94

Characterization, expression in Streptomyces lividans, and processing of the amylase of Streptomyces griseus IMRU 3570: two different amylases are derived from the same gene by an intracellular processing mechanism

alpha-Amylase gene of Streptomyces limosus: nucleotide sequence, expression motifs, and amino acid sequence homology to mammalian and invertebrate alpha-amylases

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