Peptide utilization by Pseudomonas putida and Pseudomonas maltophilia.

Pseudomonas putida assimilates peptides and hydrolyses them with intracellular peptidases. Amino acid auxotrophs (his, trp, thr or met) grew on a variety of di- and tripeptides up to twice as slowly as with free amino acids. Pseudomonas putida has separate uptake systems for both dipeptides and oligopeptides (three or more residues). Although the dipeptide system transported a variety of structurally diverse dipeptides it did not transport peptides having either unprotonatable N-terminal amino groups, blocked C-terminal carboxyl groups, D-residues, three or more residues, N-methylated peptide bonds, or beta-amino acids. Oligopeptide uptake lacked amino acid side-chain specificity, required a free N-terminal L-residue and had an upper size limit. Glycylglycyl-D,L-p-fluorophenylalanine inhibited growth of P. putida. Uptake of glycylglycyl[I-14C]alanine was rapid and inhibited by 2,4-dinitrophenol. Both dipeptide and oligopeptide uptake were constitutive. Dipeptides competed with oligopeptides for oligopeptide uptake, but oligopeptides did not compete in the dipeptide system. Final bacterial yields were 5 to 10 times greater when P. putida his was grown on histidyl di- or tripeptides rather than on free histidine because the histidyl residue was protected from catabolism by L-histidine ammonia-lyase. Methionine peptides could satisfy the methionine requirements of P. maltophilia. Generation times on glycylmethionine and glycylmethionylglycine were equal to those obtained with free methionine. Methionylglycylmethionylmethionine gave a generation time twice that of free methionine. Growth of P. maltophilia was inhibited by glycylglycyl-D,L-p-fluorophenylalanine.     

Digestion of algin by Pseudomonas maltophilia and Pseudomonas putida.

Pseudomonas maltophilia and Pseudomonas putida were identified as alginolytic species. Two media used for demonstrating alginolytic activity are described. The applied aspects of the ability of these two species to digest algin are discussed.     

嗜麦芽假单胞菌（P.maltophila）质粒的研究

本文用4种不同的方法对18株嗜麦芽假单胞菌是否含有质粒进行了检测,实验结果表明其中5株含有质粒。对其中的P2株不同生长期质粒存在状况的研究表明,该菌所含质粒与宿主不同生长期具有明显的相关性,其最高含量出现在稳定生长期,当细菌进入衰亡期时,其质粒量也随之减少,直至完全消失。 通过双向琼脂糖凝胶电泳、限制酶切分析以及分子量测定等方法,确定了嗜麦芽假单胞菌P2只含有1种质粒,分子量约为4.4×10~6道尔顿。有BamHⅠ、PitⅠ、XbaⅠ、EcoRⅠ和HindⅢ单一酶切位点。该质粒可望进一步改建成十分有用的克隆载体。     

Intracellular peptide hydrolysis by Pseudomonas putida and Pseudomonas maltophilia.

Amino acids liberated by peptidase hydrolysis of di- and oligopeptides by Pseudomonas putida were measured by trinitrobenzenesulphonate assay and high voltage electrophoresis or paper chromatography followed by ninhydrin spray. Intact bacteria or periplasmic contents released by lysozyme treatment did not hydrolyse peptides. Subcellular fractionation showed that glycylmethionine peptidase activity was cytoplasmic. This enzyme had a Km of 2 mM, and was stimulated fivefold by I mM-Co2+. Crude peptidase extract did not cleave peptides with D-residues, acylated N-terminal amino groups or N-methylated peptide bonds but otherwise showed a wide specificity. Di- or tripeptides with blocked C-terminus were hydrolysed. Leucylleucine (12 mM) and leucylglycylglycine (10 mM) did not compete with glycylmethionine (1-2 mM) and glycylmethionylglycine (1-0 mM), respectively, for hydrolysis. Pseudomonas maltophilia also contained peptidase activity (0-84 mumol amino acid released from glycylmethionylglycine/min/mg protein). Peptidases of both P. putida and P. maltophilia were constitutive.     

嗜麦芽假单胞菌黑色素的抗氧化作用研究

采用ＮＢＴ光化学反应法测定嗜麦芽假单胞菌黑色素对超氧阴离子自由基的清除作用,结果表明黑色素能明显地清除超氧阴离子自由基。４μｇ的黑色素对超氧阴离子自由基的清除率为８３．６％。黑色素有明显的抗脂质过氧化作用,能抑制鼠肝匀浆在３７℃温育下生成丙二醛,７２μｇ抑制率为９２．５％。黑色素能够降低由Ｈ２Ｏ２所致的红细胞溶血作用,经Ｈ２Ｏ２作用后,溶血率为６５．３％,经黑色素抗氧化作用后溶血率为５２．５％。     

Analysis of the lipopolysaccharide of Pseudomonas maltophilia 555

The phenol phase soluble lipopolysaccharide of Pseudomonas maltophilia strain 555, obtained from cells by the hot aqueous phenol method, was of the smooth type. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, hydrolysis, methylation, and 13C and 1H nuclear magnetic resonance analyses showed that this lipopolysaccharide has an O-chain polysaccharide composed of a repeating pentasaccharide unit, containing D-rhamnose (D-Rha, one part), 3-acetamido-3,6-dideoxy-D-galactose (D-Fuc3NAc, one part), and 4-acetamido-4,6-dideoxy-D-mannose (D-Rha4NAc, three parts) and having the structure (formula; see text) The serological cross-reactions between P. maltophilia 555 and Brucella species can now be related to the occurrence of N-acyl derivatives of 4-amino-4,6-dideoxy-D-mannopyranosyl residues in the O-chains of their respective lipopolysaccharide components.     

Characterization of Pseudomonas maltophilia Isolates from Fleece Rot

A total of 286 isolates of Pseudomonas maltophilia was collected from sheep exhibiting brown or yellowish fleece rot and from fly-strike lesions. Enzyme activities for 10 of the isolates were examined by plate tests and with the API ZYM system and compared with the enzymatic profile of a human type strain of P. maltophilia. The fact that ovine isolates of P. maltophilia are biochemically similar to pathogenic human strains suggests there may be an association between this organism and the brown to yellow type of fleece rot.     

Production of an extracellular ribonuclease by Pseudomonas maltophilia.

As part of a screening program for pseudomonad enzymes having an industrial interest, we selected ribonuclease (RNase) producing strains. Of the 150 pseudomonads screened, 6 were found to produce an extracellular RNase activity when grown on solid medium. In broth culture, the RNase activity from these six species remained bound to the cells unless gelatin was added to the medium. Gelatin was essential for the release of RNase in the broth culture, but the pH of the medium, addition of potential inducers such as nucleic acids, or addition of cations did not affect this release. However, gelatin did not appear to induce the synthesis of the enzyme. Strain B-88, identified as Pseudomonas maltophilia, was selected for further study of the enzyme. The extracellular RNase isolated from B-88 broth cultures could be separated in two fractions on the basis of the molecular weight by the ultrafiltration technique. The low molecular weight fraction reacts optimally at temperatures between 55 and 60 degrees C and optimal pH values varying from 7.4 to 9.5. At neutral or alkaline pH, the enzyme was stable at temperatures below 37 degrees C but was inactivated at 55 degrees C. The RNase was inhibited by mercury and cobalt and stimulated by magnesium.     

Induction and Catabolite Repression of Glucosidases in Pseudomonas maltophilia

Intracellular α-and β-glucosidases were induced in cell suspensions of Pseu-domonas maltophilia by maltose or cellobiose, and the synthesis of these enzymes was sensitive to apparent catabolite repression by α-ketoglutarate.     

Occurrence of Branched-Chain Hydroxy Fatty Acids in Pseudomonas maltophilia

Three branched-chain hydroxy acids not previously reported in other bacteria were found in extracts from saponified whole cells of Pseudomonas maltophilia. On the basis of evidence from mass spectrometry, infrared spectroscopy, and gas chromatographic procedures, they were identified as 2-hydroxy-9-methyldecanoic acid, 3-hydroxy-9-methyldecanoic acid, and 3-hydroxy-11-methyldodecanoic acid. These acids appeared to be tightly bound to other cellular components since they were not extracted from lyophilized cells with a chloroform-methanol (3:1) mixture.     

细菌转化黑色素的抗流感病毒作用

采用敏感的ＭＴＴ法测定了嗜麦芽假单胞菌转化黑色素的抗流感病毒作用。测定结果表明：纯化的黑色素毒性极低,对ＭＤＣＫ细胞的无毒界限为０．２ｍｇ／ｍｌ,远高于其有效的作用浓度;１０～２０μｇ／ｍｌ的黑色素能有效抑制流感病毒（血凝效价１∶３２）致细胞病变。病毒感染７２ｈ后用ＭＴＴ法测定,其宿主ＭＤＣＫ细胞保护百分率达９５％以上;此外,加黑色素后的ＭＤＣＫ细胞,抗流感病毒感染的能力优于目前抗病毒的有效药物病毒唑,前者的最佳作用浓度（２０μｇ／ｍｌ）比后者（１００μｇ／ｍｌ）低５倍。可以认为嗜麦芽假单胞菌转化黑色素具有毒性低、抗流感病毒能力强的特点     

Observations on the cell-wall compositions of the bracket fungi Laetiporus sulphureus and Piptoporus betulinus

Homogenized tissues and their alkali-soluble and alkali-insoluble fractions of fruiting bodies of the basidiomycetes Laetiporus sulphureus and Piptoporus betulinus were investigated using X-ray diffraction, infrared spectrometry and chemical methods. The presence of (13)--d-glucan, (13)--d-glucan and chitin was established. The relative amounts of these polysaccharides were different in the two species and differences were also found between context and trama. The proportion of (13)--d-glucan was exceptionally high in the context of L. sulphureus (about 78%). In addition, the trama of both species contained a substance resembling a cyclic wax by its X-ray pattern and solubility properties. The substances identified are considered to belong to the hyphal wall     

Bioproduction of low-pigment xanthan gum by a cell-wall deficient mutant of Xanthomonas campestris

Abstract

This work aims to enhance the bioproduction of xanthan gum by screening a hyper-yield producer from the wild-type Xanthomonas campestris during a long-term continuous subculture. We reported a cell-wall deficient mutant, which performed a shift of cell morphology from rod-shaped to round-shaped. Both the yield of xanthan gum and the conversion rate of feedstock were assessed using sucrose as a carbon source with the supplement of yeast extract powder, l-glutamic acid, and other raw materials. After 96?h aerobic fermentation, the yield of xanthan gum of the mutant reached up to 32?g/L, which was 3.4 times of that of the wild-type strain. The conversion rate of feedstock in the mutant was up to 92.1%, which was 3 times of that of the wild-type (31.2%). Furthermore, pigments generated were determined and compared. As a result, the fermentation broth of the wild-type performed an OD_560nm of 0.296, which was 5.8 times of that (OD_560nm?=?0.051) of the mutant. Microscopy analysis showed that the percentage of free-living cells in broth affected the color of the final product. Moreover, the robustness of the fermentation performance of the cell-wall deficient mutant at a pilot scale showed potential for industrial application.     

Demonstration and partial characterization of ADP-ribosylation in Pseudomonas maltophilia.

ADP-ribosylation of proteins occurs in many eukaryotes, and it is also the mechanism of action of a growing number of important bacterial toxins. To date, however, there is only one well-characterized ADP-ribosylation system where the ADP-ribosyltransferase and the substrate protein are both bacterial in origin, namely within the nitrogen-fixing bacterium Rhodospirillum rubrum. The present paper demonstrates the endogenous ADP-ribosylation of two proteins of Mr 32,000 and 20,000 within Pseudomonas maltophilia, a Gram-negative aerobe. The proteins have been partially purified: two apparently separate species of modified protein can be separated by ion-exchange chromatography and gel filtration (V0 and Mr 158,000 - Vi). The substrate protein(s) either has, or is co-eluted with, NAD+ glycohydrolase activity. The modification is mono-ADP-ribosyl in nature. The linkage between the acceptor amino acid and the ADP-ribose moiety is alkali-labile and stable to hydroxylamine, possibly indicating an S-glycosidic bond. The activity appears to be a true ADP-ribosylation reaction and not an NAD+ glycohydrolase activity followed by non-enzymic addition of ADP-ribose to protein. The results presented here indicate that ADP-ribosylation may have a wider significance within prokaryotic systems than previously thought.     

Expression of the mel gene from Pseudomonas maltophilia in Bacillus thuringiensis

AIMS: The objective of this work was to express a novel mel gene, responsible for melanin formation, in Bacillus thuringiensis. METHODS AND RESULTS: A novel mel gene from Pseudomonas maltophilia was sub-cloned into B. thuringiensis using a shuttle vector plasmid and electroporation. Results revealed that the mel gene was expressed under the control of the CryIIIA promoter in B. thuringiensis and conferred u.v. protection on the recipient strain. CONCLUSIONS: The novel mel gene from Ps. maltophilia expressed in B. thuringiensis conferred u.v. protection on the recipient strain. SIGNIFICANCE AND IMPACT OF THE STUDY: Products containing B. thuringiensis for pest control are sensitive to u.v. degradation. As melanin has the ability to act as a u.v. absorber, a recombinant B. thuringiensis strain producing melanin provides a new stability for B. thuringiensis preparations.