Cloning of a Serine Protease Gene From Bacillus Amyloliquefaciens and Its Expression in Bacillus Subtilis

Vasantha, N.; Rhodes, Craig; Thompson, Leo D.; Banner, Carl

doi:10.1016/b978-0-12-274160-9.50018-7

Cited by 16 publications

(15 citation statements)

References 11 publications

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“…Following a possible ribosomebinding site sequence (GGAGG, nucleotides 422 -426) a longer ORF started with a GUG codon at nucleotide 435. Although the NH2 terminus of the primary translation product of the gene is not yet known, protein synthesis should be initiated at the GUG codon as in the case of the spoVG gene from B. subtilis [37], the genes for alkaline and neutral proteases from B. amyloliquefaciens [38], and the a-amylase gene from B. stearothermophilus [32]. The NH2-terminal amino acid sequence of the B. cereus sphingomyelinase, shown in Table 2, indicates that the sphingomyelinase is synthesized in B. cereus as a secretory precursor with a signal peptide of 27 amino acid residues, and that the secretory precursor is processed at a specific cleavage site between Ala-Glu residues, resulting in the formation of the mature sphingomyelinase, which consists of 306 amino acid residues with a molecular mass of 34233 Da.…”

Section: Resultsmentioning

confidence: 99%

Nucleotide sequence and expression in Escherichia coli of the gene coding for sphingomyelinase of Bacillus cereus

Yamada

Tsukagoshi

Udaka

et al. 1988

European Journal of Biochemistry

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Bacillus cereus secretes phospholipases C , which hydrolyze phosphatidylcholine, sphingomyelin and phosphatidylinositol. A 7.5-kb HindIII fragment of B. cereus DNA cloned into Escherichia coli, with pUC18 as a vector, directed the synthesis of the sphingomyelin-hydrolyzing phospholipase C, sphingomyelinase. Nucleotide sequence analysis of the subfragment revealed that it contained two open reading frames in tandem. The upstream truncated open reading frame corresponds to the carboxy-terminal portion of the phosphatidylcholine-hydrolyzing phospholipase C, and the downstream open reading frame to the entire translational portion of the sphingomyelinase. The two phospholipase C genes form a gene cluster.As inferred from the DNA sequence, the B. cereus sphingomyelinase has a signal peptide of 27 amino acid residues and the mature enzyme comprises 306 amino acid residues, with a molecular mass of 34233 Da. The signal peptide of the enzyme was found to be functional in protein transport across the membrane of E. coli.The enzymatic properties of the sphingomyelinase synthesized in E. coli resemble those of the donor strain sphingomyelinase. The enzymatic activity toward sphingomyelin was enhanced 20 -30-fold in the presence of MgCI2, and the adsorption of the enzyme onto erythrocyte membranes was accelerated in the presence of CaCI2.Phospholipases hydrolyze several phospholipids present in biomembranes, liposomes and micelles, and play important roles in lipid metabolism. Among them only phospholipases A2 from various snake venoms and mammalian pancreas have been extensively characterized as to their primary, secondary and tertiary structures [l -51. The gene for phospholipase B of Escherichia coli (so-called 'detergent-resistant phospholipase A', which splits two fatty acids residues from phosphatides in a random order) was cloned and sequenced [6]. Phospholipases C, which cleave the bond between the hydrophobic moiety and the polar head of phospholipids, have been demonstrated in microorganisms as well as mammalian tissues, and studied for their enzymatic properties [7 -121. Phospholipases C specific for various phospholipids have been isolated from several microorganisms, such as Pseudomonas aeruginosa [13], Staphylococcus aureus [I41 and Bacillus cereus [15]. Bacillus cereus produces extracellularly three enzymes : phospholipases C, which hydrolyze phosphatidylcholine, sphingomyelin and phosphatidylinositol. Sphingomyelinase, the sphingomyelin-hydrolyzing phospholipase C produced in B. cereus, has been shown to be different from that produced in Staphylococcus aureus in several respects, such as amino acid composition and isoelectric point.Correspondence to N. Tsukagoshi,

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Section: Resultsmentioning

confidence: 99%

Nucleotide sequence and expression in Escherichia coli of the gene coding for sphingomyelinase of Bacillus cereus

Yamada

Tsukagoshi

Udaka

et al. 1988

European Journal of Biochemistry

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“…They are numbered according to that of SBPN as is conventional, and are given in full to clear up some confusions from the published sequence data and the coordinates in the Brookhaven Protein Data Bank. The sequences given for BI168 (Stahl & Ferrari, 1984), SBAS (Yoshimoto et al, 1988), SBPN (Wells, Ferrari, Henner, Estell &Chen, 1983;Vasantha, Thompson, Rhodes, Banner, Nagle & Filpula, 1984) and SBCARL (Jacobs, Eliasson, Uhlen & Flock, 1985) are those for the corresponding genes. These differ somewhat from those reported for direct sequencing of the protein for SBAS (Kurihara et al, 1972), SBPN (Markland & Smith, 1967) and SBCARL (Smith et al, 1968); the direct sequence has not been published for SBI168.…”

Section: Homology In the Bacilli Subtilisinsmentioning

confidence: 99%

Complex between the subtilisin from a mesophilic bacterium and the Leech inhibitor eglin-C

Dauter

Betzel²,

Genov³

et al. 1991

Acta Crystallogr Sect B

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“…For the membrane-bound acyl protein penicillinase, an exo form (non-acylated) is released by proteolytic processing [24]. Proteases in B. amyloliquefaciens contain an extra pro-sequence between the signal peptide and the mature protein, which is cleaved during export [38]. Drugs affect the balance between the two forms of the penicillinase [24,28,291.…”

Section: Protein Synthesis Andprocessingmentioning

confidence: 99%

Selective acylation of membrane proteins in Acholeplasma laidlawii

1986

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In membranes of the cell-wall-less prokaryote Acholeplasma laidlawii most proteins are of the integral type. A substantial fraction of these proteins are enriched in hydrophilic amino acid residues. Approximately 20 different major as well as minor proteins were found to be covalently modified with acyl chains. The same set of proteins are acylated when cells are grown in different fatty-acid-supplemented media. In individual proteins the ratio of palmitoyl/oleoyl acyl chains was 12-14 times larger than the acyl chain ratio in polar membrane lipids.The transmembrane protein D12 has close to two acyl chains per molecule. Proteins T2 and T4=, localized in the outer and inner leaflet of the membrane, respectively, occur each as pairs with a difference in relative molecular mass within each pair of approximately 2000. Each of these proteins as well as the other acyl proteins, except the light form of T4a, has close to one acyl chain per molecule. The extent of acylation was increased for certain proteins and decreased for others by treatment with globomycin or phenethylalcohol. The relative amounts of the T2 and T4= pairs were affected by these drugs. It is concluded that the mechanism of acylation is different from that in Escherichia coli lipoprotein and Bacillus penicillinase. The mean hydrophobicity [Kyte & Doolittle (1982) J. Mol. Biol. 157, 105-1321 of the A . laidlawii acyl proteins are similar to those of other bacterial acyl proteins but significantly lower than for non-acylated integral membrane proteins, supporting an anchoring function of the acyl chains. The number of membrane acyl proteins in A. laidlawii and two other mycoplasmas are at least twice that in other bacteria.

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Cloning of a Serine Protease Gene From Bacillus Amyloliquefaciens and Its Expression in Bacillus Subtilis

Cited by 16 publications

References 11 publications

Nucleotide sequence and expression in Escherichia coli of the gene coding for sphingomyelinase of Bacillus cereus

Nucleotide sequence and expression in Escherichia coli of the gene coding for sphingomyelinase of Bacillus cereus

Complex between the subtilisin from a mesophilic bacterium and the Leech inhibitor eglin-C

Selective acylation of membrane proteins in Acholeplasma laidlawii

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