Cadaverine covalently linked to a peptidoglycan is an essential constituent of the peptidoglycan necessary for the normal growth in Selenomonas ruminantium

Cadaverine links covalently to the D-glutamic acid residue of the peptidoglycan in Selenomonas ruminantium, a strictly anaerobic Gram-negative bacterium (Kamio, Y., Itoh, Y., and Terawaki, Y. (1981) J. Bacteriol. 146, 49-53). This report clarifies a physiological function of cadaverine in this organism by using DL-alpha-difluoromethyllysine, which had previously been shown to be a selective irreversible inhibitor of lysine decarboxylase of Mycoplasma dispar (P?s?, H., MaCann, P.P., Tanskanen, R., Bey, P., and Sjoerdsma, A. (1984) Biochem. Biophys. Res. Commun. 125, 205-210). DL-alpha-Difluoromethyllysine is now shown to be a potent and irreversible inhibitor of lysine decarboxylase of S. ruminantium in vitro; however, it did not inhibit the transfer of cadaverine to the alpha-carboxyl group of the D-glutamic acid residue of the peptidoglycan. DL-alpha-Difluoromethyllysine at 5 mM markedly inhibited the growth of the bacterium and caused rapid cell lysis. Immediately before the cell lysis, almost all cells became swollen, and such cells showed a loosened envelope structure when studied by electron microscopy. The peptidoglycan prepared from the DL-alpha-difluoromethyllysine-treated cells did not have covalently linked cadaverine. The growth inhibition by DL-alpha-difluoromethyllysine was completely reversed by adding cadaverine (1 mM) to the medium. Furthermore, the exogenous cadaverine was exclusively incorporated into the peptidoglycan in the presence of DL-alpha-difluoromethyllysine (5 mM), and a normal peptidoglycan was synthesized. The cell lysis and the formation of an abnormal cell structure were completely prevented by cadaverine added to the medium. We conclude that cadaverine covalently linked to the peptidoglycan in S. ruminantium is an essential constituent of the peptidoglycan and is required for cell surface integrity and the normal growth of S. ruminantium.     

Characterization of a counterpart to Mammalian ornithine decarboxylase antizyme in prokaryotes

The degradation of mammalian ornithine decarboxylase (ODC) (EC 4.1.1.17) by 26 S proteasome, is accelerated by the ODC antizyme (AZ), a trigger protein involved in the specific degradation of eukaryotic ODC. In prokaryotes, AZ has not been found. Previously, we found that in Selenomonas ruminantium, a strictly anaerobic and Gram-negative bacterium, a drastic degradation of lysine decarboxylase (LDC; EC 4.1.1.18), which has decarboxylase activities toward both L-lysine and L-ornithine with similar K(m) values, occurs upon entry into the stationary phase of cell growth by protease together with a protein of 22 kDa (P22). Here, we show that P22 is a direct counterpart of eukaryotic AZ by the following evidence. (i) P22 synthesis is induced by putrescine but not cadaverine. (ii) P22 enhances the degradation of both mouse ODC and S. ruminantium LDC by a 26 S proteasome. (iii) S. ruminantium LDC degradation is also enhanced by mouse AZ replacing P22 in a cell-free extract from S. ruminantium. (iv) Both P22 and mouse AZ bind to S. ruminantium LDC but not to the LDC mutated in its binding site for P22 and AZ. In this report, we also show that P22 is a ribosomal protein of S. ruminantium.     

Identification of a 22-kDa protein required for the degradation of Selenomonas ruminantium lysine decarboxylase by ATP-dependent protease

In Selenomonas ruminantium, a strictly anaerobic, Gram-negative bacterium isolated from sheep rumen, a rapid degradation of lysine decarboxylase (LDC) occurred on entry into the stationary phase of cell growth. Here, we identified a 22-kDa protein as a stimulating factor for the degradation of LDC, which was catalyzed by ATP-dependent protease(s) in S. ruminantium. The purified 22-kDa protein preparation itself had no degradation activity towards LDC but it was required for the degradation of LDC by ATP-dependent proteases in a cell-free system. The 22-kDa protein had similar biochemical and biophysical characteristics to those of antizyme, the regulator for the degradation of mammalian ODC, which had been reported only in mammalian cells. From the sequencing data of the N-terminal 30 amino acid residues of the 22-kDa protein preparation, 22-kDa protein was found to be a new protein which was distinguished from antizyme. This is the first report of the presence of an antizyme-like regulator protein in a prokaryote.     

Occurrence of agmatine pathway for putrescine synthesis in Selenomonas ruminatium

Selenomonas ruminantium synthesizes cadaverine and putrescine from L-lysine and L-ornithine as the essential constituents of its peptidoglycan by a constitutive lysine/ornithine decarboxylase (LDC/ODC). S. ruminantium grew normally in the presence of the specific inhibitor for LDC/ODC, DL-alpha-difluoromethylornithine, when arginine was supplied in the medium. In this study, we discovered the presence of arginine decarboxylase (ADC), the key enzyme in agmatine pathway for putrescine synthesis, in S. ruminantium. We purified and characterized ADC and cloned its gene (adc) from S. ruminantium chromosomal DNA. ADC showed more than 60% identity with those of LDC/ODC/ADCs from Gram-positive bacteria, but no similarity to that from Gram-negative bacteria. In this study, we also cloned the aguA and aguB genes, encoding agmatine deiminase (AguA) and N-carbamoyl-putrescine amidohydrolase (AguB), both of which are involved in conversion from agmatine into putrescine. AguA and AguB were expressed in S. ruminantium. Hence, we concluded that S. ruminantium has both ornithine and agmatine pathways for the synthesis of putrescine.     

Novel characteristics of Selenomonas ruminantium lysine decarboxylase capable of decarboxylating both L-lysine and L-ornithine.

Lysine decarboxylase (LDC; EC 4.1.1.18) of Selenomonas ruminantium is a constitutive enzyme and is involved in the synthesis of cadaverine, which is an essential constituent of the peptidoglycan for normal cell growth. We purified the S. ruminantium LDC by an improved method including hydrophobic chromatography and studied the fine characteristics of the enzyme. Kinetic study of LDC showed that S. ruminantium LDC decarboxylated both L-lysine and L-ornithine with similar Km and the decarboxylase activities towards both substrates were competitively and irreversibly inhibited by DL-alpha-difluoromethylornithine, which is a specific inhibitor of ornithine decarboxylase (EC 4.1.1.17). We also showed a drastic descent of LDC activity owing to the degradation of LDC at entry into the stationary phase of cell growth.     

Evolution of a novel lysine decarboxylase in siderophore biosynthesis

Structural backbones of iron‐scavenging siderophore molecules include polyamines 1,3‐diaminopropane and 1,5‐diaminopentane (cadaverine). For the cadaverine‐based desferroxiamine E siderophore in Streptomyces coelicolor, the corresponding biosynthetic gene cluster contains an ORF encoded by desA that was suspected of producing the cadaverine (decarboxylated lysine) backbone. However, desA encodes an l ‐2,4‐diaminobutyrate decarboxylase (DABA DC) homologue and not any known form of lysine decarboxylase (LDC). The only known function of DABA DC is, together with l ‐2,4‐aminobutyrate aminotransferase (DABA AT), to synthesize 1,3‐diaminopropane. We show here that S. coelicolor desA encodes a novel LDC and we hypothesized that DABA DC homologues present in siderophore biosynthetic clusters in the absence of DABA AT ORFs would be novel LDCs. We confirmed this by correctly predicting the LDC activity of a DABA DC homologue from a Yersinia pestis siderophore biosynthetic pathway. The corollary was confirmed for a DABA DC homologue, adjacent to a DABA AT ORF in a siderophore pathway in the cyanobacterium Anabaena variabilis, which was shown to be a bona fide DABA DC. These findings enable prediction of whether a siderophore pathway will utilize 1,3‐diaminopropane or cadaverine, and suggest that the majority of bacteria use DABA AT and DABA DC for siderophore, rather than norspermidine/polyamine biosynthesis.     

Cadaverine covalently linked to the peptidoglycan serves as the correct constituent for the anchoring mechanism between the outer membrane and peptidoglycan in Selenomonas ruminantium

In Selenomonas ruminantium, a strictly anaerobic and gram-negative bacterium, cadaverine covalently linked to the peptidoglycan is required for the interaction between the peptidoglycan and the S-layer homologous (SLH) domain of the major outer membrane protein Mep45. Here, using a series of diamines with a general structure of NH(3)(+)(CH(2))(n)NH(3)(+) (n = 3 to 6), we found that cadaverine (n = 5) specifically serves as the most efficient constituent of the peptidoglycan in acquiring the high resistance of the cell to external damage agents and is required for effective interaction between the SLH domain of Mep45 and the peptidoglycan, facilitating the correct anchoring of the outer membrane to the peptidoglycan.     

Identification of the amino acid residues conferring substrate specificity upon Selenomonas ruminantium lysine decarboxylase

Lysine decarboxylase (LDC, EC 4.1.1.18) from Selenomonas ruminantium has decarboxylating activities towards both L-lysine and L-ornithine with similar K(m) and Vmax. Here, we identified four amino acid residues that confer substrate specificity upon S. ruminantium LDC and that are located in its catalytic domain. We have succeeded in converting S. ruminantium LDC to an enzyme with a preference in decarboxylating activity for L-ornithine when the four-residue of LDC were replaced by the corresponding residues of mouse ornithine decarboxylase (EC 4.1.1.17).     

Novel Characteristics of Selenomonas ruminantium Lysine Decarboxylase Capable of Decarboxylating Both L-Lysine and L-Ornithine

Lysine decarboxylase (LDC; EC 4.1.1.18) of Selenomonas ruminantium is a constitutive enzyme and is involved in the synthesis of cadaverine, which is an essential constituent of the peptidoglycan for normal cell growth. We purified the S. ruminantium LDC by an improved method including hydrophobic chromatography and studied the fine characteristics of the enzyme. Kinetic study of LDC showed that S. ruminanitum LDC decarboxylated both L-lysine and L-ornithine with similar K _m and the decarboxylase activities towards both substrates were competitively and irreversibly inhibited by DL-α-difluoromethylornithine, which is a specific inhibitor of ornithine decarboxylase (EC 4.1.1.17). We also showed a drastic descent of LDC activity owing to the degradation of LDC at entry into the stationary phase of cell growth.     

Gene cloning and molecular characterization of lysine decarboxylase from Selenomonas ruminantium delineate its evolutionary relationship to ornithine decarboxylases from eukaryotes

Lysine decarboxylase (LDC; EC 4.1.1.18) from Selenomonas ruminantium comprises two identical monomeric subunits of 43 kDa and has decarboxylating activities toward both L-lysine and L-ornithine with similar K(m) and V(max) values (Y. Takatsuka, M. Onoda, T. Sugiyama, K. Muramoto, T. Tomita, and Y. Kamio, Biosci. Biotechnol. Biochem. 62:1063-1069, 1999). Here, the LDC-encoding gene (ldc) of this bacterium was cloned and characterized. DNA sequencing analysis revealed that the amino acid sequence of S. ruminantium LDC is 35% identical to those of eukaryotic ornithine decarboxylases (ODCs; EC 4.1.1.17), including the mouse, Saccharomyces cerevisiae, Neurospora crassa, Trypanosoma brucei, and Caenorhabditis elegans enzymes. In addition, 26 amino acid residues, K69, D88, E94, D134, R154, K169, H197, D233, G235, G236, G237, F238, E274, G276, R277, Y278, K294, Y323, Y331, D332, C360, D361, D364, G387, Y389, and F397 (mouse ODC numbering), all of which are implicated in the formation of the pyridoxal phosphate-binding domain and the substrate-binding domain and in dimer stabilization with the eukaryotic ODCs, were also conserved in S. ruminantium LDC. Computer analysis of the putative secondary structure of S. ruminantium LDC showed that it is approximately 70% identical to that of mouse ODC. We identified five amino acid residues, A44, G45, V46, P54, and S322, within the LDC catalytic domain that confer decarboxylase activities toward both L-lysine and L-ornithine with a substrate specificity ratio of 0.83 (defined as the k(cat)/K(m) ratio obtained with L-ornithine relative to that obtained with L-lysine). We have succeeded in converting S. ruminantium LDC to form with a substrate specificity ratio of 58 (70 times that of wild-type LDC) by constructing a mutant protein, A44V/G45T/V46P/P54D/S322A. In this study, we also showed that G350 is a crucial residue for stabilization of the dimer in S. ruminantium LDC.     

Chemical structure of peptidoglycan in Selenomonas ruminantium: cadaverine links covalently to the D-glutamic acid residue of peptidoglycan

The peptidoglycan of Selenomonas ruminantium, a strictly anaerobic bacterium, contains cadaverine (Y. Kamio, Y. Itoh, Y. Terawaki, and T. Kusano, J. Bacteriol. 145:122-128, 1981). This report describes the chemical structure of the peptidoglycan of this bacterium. The [14C]cadaverine-labeled peptidoglycan was degraded with the lytic enzymes prepared from Streptomyces albus G into three small fragments including a major fragment (band A compound). Bank A compound was composed of L-alanine, D-glutamic acid, meso-diaminopimelic acid, D-alanine, and cadaverine in the molar ratio 0.98:1.0:1.0:0.98:0.97. Diaminopimelic acid, L-alanine, and cadaverine were N-terminal residues in band A compound. When the [14C]cadaverine-labeled band A compound was subjected to partial acid hydrolysis, two peptide fragments were obtained. One of them consisted of diaminopimelic acid and D-alanine; diaminopimelic acid was the N-terminal amino acid, and the other fragment was composed of L-alanine, D-glutamic acid, and cadaverine, of which L-alanine and cadaverine were N-terminal. These results lead us to conclude that the primary peptide structure of band A compound is L-alanyl-D-glutamyl-meso-diaminopimelyl-D-alanine and that cadaverine links covalently to the D-glutamic acid residue.     

Improved metabolic action of a bacterial lysine decarboxylase gene in tobacco hairy root cultures by its fusion to arbcS transit peptide coding sequence

The gene of a bacterial lysine decarboxylase (ldc) fused to arbcS transit peptide coding sequence (tp), and under the control of the CaMV 35S promoter, was expressed in hairy root cultures ofNicotiana tabacum. The fusion of theldc to the targeting signal sequence improved the performance of the bacterial gene in the plant cells in many respects. Nearly all transgenic hairy root cultures harbouring the35S-tp-ldc gene contained distinctly higher lysine decarboxylase activity (from 1.5 to 30 pkat LDC per mg protein) than those which had been transformed with constructs in which the gene had been directly cloned behind the CaMV 35S promoter. The higher enzyme activity led to the accumulation of up to 0.7% cadaverine on a dry mass basis. In addition, part of the cadaverine pool was used for increased biosynthesis of anabasine, an alkaloid which was hardly detectable in control cultures. The best line contained anabasine levels of 0.5% dry mass, which could further be enhanced by feeding of lysine.     

Characterization of a major envelope protein from the rumen anaerobe Selenomonas ruminantium OB268

Cell envelopes from the Gram-negative staining but phylogenetically Gram-positive rumen anaerobe Selenomonas ruminantium OB268 contained a major 42 kDa heat modifiable protein. A similarly sized protein was present in the envelopes of Selenomonas ruminantium D1 and Selenomonas infelix. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of Triton X-100 extracted cell envelopes from S. ruminantium OB268 showed that they consisted primarily of the 42 kDa protein. Polyclonal antisera produced against these envelopes cross-reacted only with the 42 kDa major envelope proteins in both S. ruminantium D1 and S. infelix, indicating a conservation of antigenic structure among each of the major envelope proteins. The N-terminus of the 42 kDa S. ruminantium OB268 envelope protein shared significant homology with the S-layer (surface) protein from Thermus thermophilus, as well as additional envelope proteins containing the cell surface binding region known as a surface layer-like homologous (SLH) domain. Thin section analysis of Triton X-100 extracted envelopes demonstrated the presence of an outer bilayer over-laying the cell wall, and a regularly ordered array was visible following freeze-fracture etching through this bilayer. These findings suggest that the regularly ordered array may be composed of the 42 kDa major envelope protein. The 42 kDa protein has similarities with regularly ordered outer membrane proteins (rOMP) reported in certain Gram-negative and ancient eubacteria.     

Cadaverine is covalently linked to peptidoglycan in Selenomonas ruminantium.

Cadaverine was found to exist as a component of cell wall peptidoglycan of Selenomonas ruminantium, a strictly anaerobic bacterium. [14C]cadaverine added to the growth medium was incorporated into the cells, and about 70% of the total radioactivity incorporated was found in the peptidoglycan fraction. When the [14C]cadaverine-labeled peptidoglycan preparation was acid hydrolyzed, all of the 14C counts were recovered as cadaverine. The [14C]cadaverine-labeled peptidoglycan preparation was digested with lysozyme into three small fragments which were radioactive and were positive in ninhydrin reaction. One major spot, a compound of the fragments, was composed of alanine, glutamic acid, diaminopimelic acid, cadaverine, muramic acid, and glucosamine. One of the two amino groups of cadaverine was covalently linked to the peptidoglycan, and the other was free. The chemical composition of the peptidoglycan preparation of this strain was determined to be as follows: L-alanine-D-alanine-D-glutamic acid-meso-diaminopimelic acid-cadaverine-muramic acid-glucosamine (1.0:1.0:1.0:1.0:1.1:0.9:1.0).     

Metabolic effects of a bacterial lysine decarboxylase gene expressed in a hairy root culture ofNicotiana glauca

OneNicotiana glauca line with distinctly enhanced levels of lysine decarboxylase (LDC) activity and of cadaverine was detected among 54 hairy root cultures of different tobacco species, transformed with the binary vector pLX222 carrying a bacteial lysine decarboxylase gene directed by the 35S-promoter of CaMV. Anabasine levels of this line were nearly doubled in comparison to control lines transformed with the gus-gene instead of the ldc-gene.     

Cadaverine Covalently Linked to Peptidoglycan Is Required for Interaction between the Peptidoglycan and the Periplasm-Exposed S-Layer-Homologous Domain of Major Outer Membrane Protein Mep45 in Selenomonas ruminantium

The peptidoglycan of Selenomonas ruminantium is covalently bound to cadaverine (PG-cadaverine), which likely plays a significant role in maintaining the integrity of the cell surface structure. The outer membrane of this bacterium contains a 45-kDa major protein (Mep45) that is a putative peptidoglycan-associated protein. In this report, we determined the nucleotide sequence of the mep45 gene and investigated the relationship between PG-cadaverine, Mep45, and the cell surface structure. Amino acid sequence analysis showed that Mep45 is comprised of an N-terminal S-layer-homologous (SLH) domain followed by α-helical coiled-coil region and a C-terminal β-strand-rich region. The N-terminal SLH domain was found to be protruding into the periplasmic space and was responsible for binding to peptidoglycan. It was determined that Mep45 binds to the peptidoglycan in a manner dependent on the presence of PG-cadaverine. Electron microscopy revealed that defective PG-cadaverine decreased the structural interactions between peptidoglycan and the outer membrane, consistent with the proposed role for PG-cadaverine. The C-terminal β-strand-rich region of Mep45 was predicted to be a membrane-bound unit of the 14-stranded β-barrel structure. Here we propose that PG-cadaverine possesses functional importance to facilitate the structural linkage between peptidoglycan and the outer membrane via specific interaction with the SLH domain of Mep45.Polyamines, the ubiquitous polycationic compounds composed of a hydrocarbon backbone with multiple amino groups, exist in all living cells and participate in a wide variety of biological reactions, including DNA, RNA, and protein synthesis (34). However, it has been revealed that some strictly anaerobic eubacteria belonging to the Veillonellaceae family, such as Selenomonas ruminantium, Veillonella alcalescens, Veillonella parvula, and Anaerovibrio lipolyticus, possess polyamines covalently linked to their peptidoglycan (PG) as an essential constituent (8, 16, 17). S. ruminantium possesses a peptidoglycan associated with cadaverine. Cadaverine binds covalently to the α-carboxyl group of the d-glutamic acid residue of peptidoglycan by one of its two amino groups, and the other amino group remains as a free cation (15). In this bacterium, cadaverine is synthesized constitutively from lysine by lysine/ornithine decarboxylase (LDC/ODC [EC 4.1.1.18]), a bifunctional enzyme that decarboxylates both l-lysine and l-ornithine at similar K_m and V_max values (35, 36) and is transferred to a d-glutamic acid residue by a particulate enzyme designated as lipid intermediate:diamine transferase (20). The cadaverine synthesis by LDC/ODC is completely inhibited by dl-α-difluoromethyllysine (DFML) or dl-α-difluoromethylornithine (DFMO), which inhibits the decarboxylating activity toward both l-lysine and l-ornithine (35), and the prevention of the cadaverine synthesis in S. ruminantium was shown to lead to the significant decrease of the amount of the cadaverine covalently linked to peptidoglycan (PG-cadaverine) and result in the growth inhibition (17). Since this inhibitory effect accompanies a drastic morphological change of the cells resulting in an aberrant cell surface structure, PG-cadaverine has been assumed to play a significant role in maintaining the integrity of the cell surface (17).The cell surface structure of S. ruminantium has a typical Gram-negative three-layer organization, comprising a cytoplasmic membrane, peptidoglycan layer, and outer membrane (18). However, it contains neither the free nor bound form of murein-lipoprotein (19), which plays an important role in the structural linkage between the outer membrane and peptidoglycan, thereby maintaining the structural integrity of the cell surface structures of Gram-negative bacteria (5, 33). The Escherichia coli lpo mutant that lacks murein-lipoprotein becomes hypersensitive to EDTA, resulting in rapid cell lysis upon exposure to EDTA. In contrast, S. ruminantium shows no cell lysis, even in the presence of high concentrations of EDTA, despite the absence of murein-lipoprotein (19). One possible interpretation for these findings was the assumption that PG-cadaverine associates with the structural connection between the outer membrane and peptidoglycan, thereby replacing the function of murein-lipoprotein with an outer membrane component or components. Nevertheless, the factors in the outer membrane interacting with PG-cadaverine have not been identified.The outer membrane of S. ruminantium contains a 45-kDa major protein (Mep45), which has been proposed to be a peptidoglycan-associating protein (18, 19). Kalmokoff et al. reported that the major outer membrane protein of S. ruminantium OB268, which is similar to Mep45 in size, contains an N-terminal surface-layer homology (SLH) domain (13), a putative functional domain that interacts with cell wall components (27). These findings prompted us to investigate the Mep45 major outer membrane protein of S. ruminantium as a putative outer membrane component interacting with PG-cadaverine. In this report, we characterize the Mep45 protein and its interactions with PG-cadaverine and prove their involvement in the structural linkage between the outer membrane and peptidoglycan.     

Regulation of cadaverine and putrescine levels in different organs of chick-pea seed and seedlings during germination

In chick-pea ( Cicer arietinum L.) seed germinated in the presence of ¹⁴C-lysine, the latter is taken up and partly metabolised to cadaverine and TCA-precipitable molecules. Labelled cadaverine is detectable in seedlings only after 3 days, on a labelled lysine-containing medium, as confirmed also by the presence of lysine decarboxylase (LDC) activity, measured in the embryo axis and cotyledons of the seed and in the epicotyl, cotyledons, hypocotyl and roots of the seedling on the basis of ¹⁴CO₂ evolution from the labelled precursor. Putrescine biosynthesis occurred only via arginine decarboxylase (ADC) activities in soaked seeds and via both ADC and ornithine decarboxylase (ODC) activities in seedlings. Both putrescine and cadaverine were present in soaked seed, and accumulated in very large amounts in the different portions of both 3- and 8-day-old seedlings, while spermidine and spermine titers were maintained at similar levels with respect to the seed. Diamine oxidase activity, measured by evaluating oxygen consumption in the presence of putrescine, was absent in ungerminated seed and appeared in 3- and 8-day-old seedlings. In order to clarify the metabolic relationships between cadaverine and the more common polyamines, gradients of biosynthesis, accumulation and degradation of putrescine and cadaverine along the seedling axis were compared, indicating that the two diamines behave similarly during seed germination and seedling development. Their conspicuous accumulation (up to 6 m M for putrescine) seems to be regulated mainly via oxidation rather than biosynthesis.     

Increased production of cadaverine and anabasine in hairy root cultures of Nicotiana tabacum expressing a bacterial lysine decarboxylase gene

Several hairy root cultures of Nicotiana tabacum varieties, carrying two direct repeats of a bacterial lysine decarboxylase (ldc) gene controlled by the cauliflower mosaic virus (CaMV) 35S promoter expressed LDC activity up to 1 pkat/mg protein. Such activity was, for example, sufficient to increase cadaverine levels of the best line SR3/1-K1,2 from ca. 50 g (control cultures) to about 700 g/g dry mass. Some of the overproduced cadaverine of this line was used for the formation of anabasine, as shown by a 3-fold increase of this alkaloid. In transgenic lines with lower LDC activity the changes of cadaverine and anabasine levels were correspondingly lower and sometimes hardly distinguishable from controls. Feeding of lysine to root cultures, even to those with low LDC activity, greatly enhanced cadaverine and anabasine livels, while the amino acid had no or very little effect on controls and LDC-negative lines.     

Structural specificity of diamines covalently linked to peptidoglycan for cell growth of Veillonella alcalescens and Selenomonas ruminantium.

Putrescine and cadaverine are essential constituents of the peptidoglycan of Veillonella alcalescens, Veillonella parvula, and Selenomonas ruminantium and are necessary for the growth of these organisms (Y. Kamio and K. Nakamura, J. Bacteriol. 169:2881-2884, 1987, and Y. Kamio, H. P?s?, Y. Terawaki, and L. Paulin, J. Biol. Chem. 261:6585-6589, 1986). In this study, the structural specificity of the diamine requirement for normal cell growth of these bacteria was examined by using a series of diamines with a general structure of NH3+ X (CH2)n X NH3+. Diaminohexane (n = 6) which was incorporated into the peptidoglycan was as effective as putrescine (n = 4) and cadaverine (n = 5) for normal cell growth. However, diaminopropane (n = 3) and diaminoheptane (n = 7) were less effective for growth than diaminohexane, although they were incorporated into the peptidoglycan to the same extent.