Antibodies against Mucin‐Based Glycopeptides Affect Trypanosoma cruzi Cell Invasion and Tumor Cell Viability

This study describes the synthesis of glycopeptides NHAc[βGal]‐(Thr)₂‐[αGalNAc]‐(Thr)₂‐[αGlcNAc]‐(Thr)₂Gly‐OVA ( 1 ‐OVA) and NHAc[βGal‐αGalNAc]‐(Thr)₃‐[αLacNAc]‐(Thr)₃‐Gly‐OVA ( 2 ‐OVA) as mimetics of both T. cruzi and tumor mucin glycoproteins. These glycopeptides were obtained by solid‐phase synthesis, which involved the prior preparation of the protected glycosyl amino acids αGlcNAc‐ThrOH ( 3 ), αGalNAc‐ThrOH ( 4 ), βGal‐ThrOH ( 5 ), αLacNAc‐ThrOH ( 6 ), and βGal‐αGalNAc‐ThrOH ( 7 ) through glycosylation reactions. Immunizations of mice with glycopeptides 1 ‐OVA and 2 ‐OVA induced high antibody titers (1:16 000), as verified by ELISA tests, whereas flow cytometry assays showed the capacity of the obtained anti‐glycopeptides 1 ‐OVA and 2 ‐OVA antibodies to recognize both T. cruzi and MCF‐7 tumor cells. In addition, antisera induced by glycopeptides 1 ‐OVA and 2 ‐OVA were also able to inhibit T. cruzi fibroblast cell invasion (70 %) and to induce antibody‐mediated cellular cytotoxicity (ADCC) against MCF‐7 cells, with 50 % reduction of cell viability.     

The inverse type II β-turn on D-Trp-Phe, a pharmacophoric motif for MOR agonists

Herein we propose the D ‐Trp‐Phe sequence within an inverse type II β‐turn as a new kind of pharmacophoric motif for μ‐opioid receptor (MOR) cyclopeptide agonists. Initially, we observed that c[Tyr‐D ‐Pro‐D ‐Trp‐Phe‐Gly] ( 4 ), an analogue of endomorphin‐1 (H‐Tyr‐Pro‐Trp‐Phe‐NH₂) lacking the crucial protonatable amino group of Tyr 1, is a MOR agonist with 10^?8 M affinity. Molecular docking analysis suggested that the relevant interactions with the receptor involve D ‐Trp‐Phe. The bioactive conformation of this region was investigated by selected derivatives of 4 designed to adopt an inverse type II β‐turn. These efforts led to c[Tyr‐Gly‐D ‐Trp‐Phe‐Gly] ( 14 ) and to the cyclotetrapeptide c[D ‐Asp‐1‐amide‐β‐Ala‐D ‐Trp‐Phe] ( 15 ), showing improved nanomolar affinity. Both 14 and 15 selectively bind MOR, as they have negligible affinity for the κ‐ and δ‐opioid receptors. Both 14 and 15 behave as partial MOR agonists in functional assays. Conformational and docking analyses confirm the role of the inverse β‐turn in binding. These results indicate that the D ‐Trp‐Phe inverse β‐turn structure can be used for designing non‐endomorphin‐like peptidomimetic opioid agonists in general, characterized by an atypical mechanism of interaction between ligand and receptor.     

Model polypeptide of mussel adhesive protein. I. Synthesis and adhesive studies of sequential polypeptides (X‐Tyr‐Lys)n and (Y‐Lys)n

The sequential polytripeptides and polydipeptides, (X‐Tyr‐Lys)_n, (XGly, Ala, Pro, Ser, Leu, Ile, Phe), (Y‐Lys)_n, (YGly, Tyr), and (Gly‐Tyr)_n, which imitate a mussel adhesive protein, have been synthesized. The molecular weights of the polypeptides were estimated to be 7,200 ∼ 13,400 (19 ∼ 42 repeating units), and the polypeptides were found to have satisfactory amino acid sequences. The polypeptides were crosslinked by tyrosinase, and the optimal pH in the crosslinking reaction was 7.4 in the case of the polytripeptide, (Gly‐Tyr‐Lys)_n. The optimal tyrosinase amount for the adhesive strength of (Gly‐Tyr‐Lys)_n was 0.34 unit/mg (polypeptide) at pH 7.4. The shear adhesive strength of the polytripeptide increased with an increase in the polytripeptide concentration, and was not influenced by the third amino acid, X. The shear adhesive strengths of polytripeptides (X‐Tyr‐Lys)_n were equal to one of the synthetic polydecapeptides, (Ala‐Lys‐Pro‐Ser‐Tyr‐Pro‐Pro‐Thr‐Tyr‐Lys)_n and (Gly‐Pro‐Lys‐Thr‐Tyr‐Pro‐Pro‐Thr‐Tyr‐Lys)_n which were the model polydecapeptides for blue mussel and Californian mussel, respectively. © 2000 John Wiley & Sons, Inc. J Appl Polym Sci 76: 929–937, 2000     

An Engineered Biocatalyst for the Synthesis of Glycoconjugates: Utilization of β1,3‐N‐Acetyl‐D‐glucosaminyltransferase from Streptococcus agalactiae Type Ia Expressed in Escherichia coli as a Fusion with Maltose‐Binding Protein

A fusion protein composed of β1,3‐N‐acetyl‐D ‐glucosaminyltransferase (β1,3‐GlcNAcT) from Streptococcus agalactiae type Ia and maltose‐binding protein (MBP) was produced in Escherichia coli as a soluble and highly active form. Although this fusion protein (MBP‐β1,3‐GlcNAcT) did not show any sugar‐elongation activity to some simple low‐molecular weight acceptor substrates such as galactose, Galβ(1→4)Glc (lactose), Galβ(1→4)GlcNAc (N‐acetyllactosamine), Galβ(1→4)GlcNAcβ(1→3)Galβ(1→4)Glc (lacto‐N‐tetraose), and Galβ(1→4)GlcβCer (lactosylceramide, LacCer), the multivalent glycopolymer having LacCer‐mimic branches (LacCer mimic polymer, LacCer primer) was found to be an excellent acceptor substrate for the introduction of a β‐GlcNAc residue at the O‐3 position of the non‐reducing galactose moiety by this engineered enzyme. Subsequently, the polymer having GlcNAcβ(1→3)Galβ(1→4)Glc was subjected to further enzymatic modifications by using recombinant β1,4‐D ‐galactosyltransferase (β1,4‐GalT), α2,3‐sialyltransferase (α2,3‐SiaT), α1,3‐L ‐fucosyltransferase (α1,3‐FucT), and ceramide glycanase (CGase) to afford a biologically important ganglioside; Neu5Aα(2→3)Galβ(1→4)[Fucα(1→3)]GlcNAcβ(1→3)Galβ(1→4)GlcCerα(IV3Neu5Acα,III3Fucα‐nLc4Cer) in 40% yield (4 steps). Interestingly, it was suggested that MBP‐β1,3‐GlcNAcT could also catalyze a glycosylation reaction of the LacCer mimic polymer with N‐acetyl‐D ‐galactosamine served from UDP‐GalNAc to afford a polymer carrying trisaccharide branches, GalNAcβ(1→3)Galβ(1→4)Glc. The versatility of the MBP‐β1,3‐GlcNAcT in the practical synthesis was preliminarily demonstrated by applying this fusion protein as an immobilized biocatalyst displayed on the amylose resin which is known as a solid support showing potent binding‐affinity with MBP.     

Indoloazepinone‐Constrained Oligomers as Cell‐Penetrating and Blood–Brain‐Barrier‐Permeating Compounds

Non‐cationic and amphipathic indoloazepinone‐constrained (Aia) oligomers have been synthesized as new vectors for intracellular delivery. The conformational preferences of the [l ‐Aia‐Xxx]_n oligomers were investigated by circular dichroism (CD) and NMR spectroscopy. Whereas Boc‐[l ‐Aia‐Gly]_2,4‐OBn oligomers 12 and 13 and Boc‐[l ‐Aia‐β³‐h‐l ‐Ala]_2,4‐OBn oligomers 16 and 17 were totally or partially disordered, Boc‐[l ‐Aia‐l ‐Ala]₂‐OBn ( 14 ) induced a typical turn stabilized by C₅‐ and C₇‐membered H‐bond pseudo‐cycles and aromatic interactions. Boc‐[l ‐Aia‐l ‐Ala]₄‐OBn ( 15 ) exhibited a unique structure with remarkable T‐shaped π‐stacking interactions involving the indole rings of the four l ‐Aia residues forming a dense hydrophobic cluster. All of the proposed FITC‐6‐Ahx‐[l ‐Aia‐Xxx]₄‐NH₂ oligomers 19 – 23 , with the exception of FITC‐6‐Ahx‐[l ‐Aia‐Gly]₄‐NH₂ ( 18 ), were internalized by MDA‐MB‐231 cells with higher efficiency than the positive references penetratin and Arg₈. In parallel, the compounds of this series were successfully explored in an in vitro blood–brain barrier (BBB) permeation assay. Although no passive diffusion permeability was observed for any of the tested Ac‐[l ‐Aia‐Xxx]₄‐NH₂ oligomers in the PAMPA model, Ac‐[l ‐Aia‐l ‐Arg]₄‐NH₂ ( 26 ) showed significant permeation in the in vitro cell‐based human model of the BBB, suggesting an active mechanism of cell penetration.     

Mutations to alter Aspergillus awamori glucoamylase selectivity. III. Asn20-->Cys/Ala27-->Cys, Ala27-->Pro, Ser30-->Pro, Lys108-->Arg, Lys108- ->Met, Gly137-->Ala, 311-314 Loop, Tyr312-->Trp and Ser436-->Pro

Mutations Asn20-->Cys/Ala27-->Cys (SS), Ala27-->Pro, Ser30-->Pro, Lys108-->Arg, Gly137-->Ala, Tyr312-->Trp and Ser436-->Pro in Aspergillus awamori glucoamylase, along with a mutation inserting a seven-residue loop between Tyr311 and Gly314 (311-314 Loop), were made to increase glucose yield from maltodextrin hydrolysis. No active Lys108-->Met glucoamylase was found in the supernatant after being expressed from yeast. Lys108-->Arg, 311-314 Loop and Tyr312-->Trp glucoamylases have lower activities than wild-type glucoamylase; other GAs have the same or higher activities. SS and 311-314 Loop glucoamylases give one-quarter to two-thirds the relative rates of isomaltose formation from glucose compared with glucose formation from maltodextrins at 35, 45 and 55 degrees C, correlating with up to 2% higher peak glucose yields from 30% (w/v) maltodextrin hydrolysis. Conversely, Lys108-->Arg glucoamylase has relative isomaltose formation rates three times higher and glucose yields up to 4% lower than wild- type glucoamylase. Gly137-->Ala and Tyr312-->Trp glucoamylases also give high glucose yields at higher temperatures. Mutated glucoamylases that catalyze high rates of isomaltose formation give higher glucose yields from shorter than from longer maltodextrins, opposite to normal experience with more efficient glucoamylases.      

Random mutagenesis into the conserved Gly154 of subtilisin E: isolation and characterization of the revertant enzymes

We analyzed the role played by the conserved Gly154, a constituent of the P1 substrate-binding pocket of Bacillus subtilis subtilisin E, in the catalytic properties of the protease. Using an Escherichia coli expression system, the termination codon at position 154 in subtilisin E was first introduced to abolish the catalytic activity through truncation of the C-terminus from amino acid residues 154-275. We then attempted to obtain revertants with substitutions of various amino acids at position 154 by the polymerase chain reaction using a mixture of oligonucleotides. In addition to the Gly residue (wild-type), six amino acid substitutions (Ala, Arg, Leu, Phe, Pro and Thr) gave caseinolytic activity. When assayed with synthetic peptide substrates, most of the revertants showed a considerable decrease in specific activity and a P1 specificity similar to that of the wild-type enzyme. An Ala154 mutant purified from the periplasmic space in E. coli, however, resulted in an up to 2.3-fold preference for Val rather than Pro as a P2 substrate relative to the wild-type. Further, a significant 2-10-fold increase in the catalytic efficiency occurred in the Gly127Ala plus Gly154Ala combination variant, relative to the single Gly127Ala variant, without any change in the restricted specificity. The kinetic data and molecular modeling analysis demonstrate the important role of position 154 in the catalytic efficiency as well as in the substrate specificity of subtilisin E.      

Vero细胞批式培养中的氨基酸代谢

用高效液相色谱(HPLC)测定了批式培养的Vero细胞培养液中氨基酸的变化,结果表明Vero细胞在生长过程中需要消耗12种氨基酸,同时生成4种氨基酸, Vero细胞在生长期吸收Leu,Thr,Ile,Lys,Phe和Tyr较多,而在静止期这些氨基酸却消耗较少.     

1,6‐hexanediol diacrylate‐crosslinked polystyrene: Preparation,characterization, and application in peptide synthesis

A copolymer of 1,6‐hexanediol diacrylate (HDODA) and styrene was prepared by a suspension polymerization method. The resin was characterized by infrared and carbon‐13 cross‐polarization magic‐angle spin (¹³C CP‐MAS) spectroscopy. The topology of the resin was examined by scanning electron microscopy (SEM). The polymer swells extensively in common solvents used for peptide synthesis. The resin exhibited chemical stability even in neat trifluoroacetic acid. The applicability of the new resin was demonstrated by synthesis of Val‐Ala‐Val‐Ala‐Ala‐Gly, Gln‐Val‐Gly‐Gln‐Val‐Glu‐Leu‐Gly, and Val‐Gln‐Ala‐Ala‐Ile‐Asp‐Tyr‐Ile‐Asn‐Gly. Comparative synthetic studies showed that the new resin is superior to divinylbenzene (DVB)‐based resin in the case of the synthesis of hydrophobic peptide sequences. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 87: 1290–1296, 2003     

Use of fortuitous in vitro mutations in a synthetic Cu-Zn superoxide dismutase gene to illuminate protein structure-function relationships

A completely synthetic bovine copper-zinc superoxide dismutase gene (Cu- ZnSOD), designed using the most favoured codons for expression in yeast, was constructed. Fortuitous mutations introduced while cloning the synthetic gene permitted the additional construction of four altered-polypeptide products representing two single (Pro121-->Leu and Gly128-->Asp), one double (Pro100-->Leu, Arg113-->Lys) and one triple (Pro100-->Leu, Arg113-->Lys, Pro121-->Leu) mutant. All five versions of the gene were expressed in a SOD-deficient Escherichia coli strain. The 'wild-type' version of the gene and the two single-mutants were expressed to equal extents (approximately 8% of total soluble protein). However, compared with the 'wild-type' enzyme, one single-mutant (Gly128-->Asp) showed almost twice as much dismutase activity whilst the other (Pro121-->Leu) exhibited only 70% of the 'wild-type' level. In contrast, the double and triple mutants showed diminished expression of the gene (approximately 1 and 3% of total soluble protein, respectively) and almost no detectable SOD activity. Polyclonal antibovine SOD antibody bound all the recombinant proteins, although some of the products showed decreased size and probably altered conformations. The 'wild-type' superoxide dismutase recombinant was correctly dimerized and possessed dismutase activity, as did the Gly128- ->Asp mutant despite the change in charge. Mutations in the other three versions affected enzyme folding and activity. The effect of the different mutations appeared to be additive, with the Pro121-->Leu substitution leading to the apparent proteolytic degradation of the enzyme in vivo.      

Optical Resolution Membranes from Polysulfones Bearing Alanine Derivatives as Chiral Selectors

Polysulfones bearing a derivative of alanyl residue employed as chiral selectors were prepared by polymer modification. The specific rotation ([α]_D) of the polysulfone with a derivative of D ‐alanyl residue (PSf‐Ac‐D ‐Ala) was determined to be 2.87 deg · cm² · g^?1 (c = 1.00 g · dL^?1 in DMF) and that with L ‐alanyl residue (PSf‐Ac‐L ‐Ala) to be ‐2.36 deg · cm² · g^?1 (c = 1.00 g · dL^?1 in DMF). The membrane from PSf‐Ac‐D ‐Ala preferentially adsorbed the D ‐isomer of Glu from racemic mixture of Glu and vice versa. Chiral separation ability was studied by applying a potential difference as a driving force for membrane transport. The permselectivity of PSf‐Ac‐D ‐Ala toward D ‐Glu (α_D/L) was determined to be 1.40, and that of PSf‐Ac‐L ‐Ala toward the L ‐isomer (α_L/D) to be 1.48 at 18.0 V, reflecting their adsorption selectivity.

     

A Thioether‐Stabilized d‐Proline–l‐Proline‐Induced β‐Hairpin Peptide of Defensin Segment Increases Its Anti‐Candida albicans Ability

We report a β‐hairpin dual stabilizing strategy: a d ‐proline‐l ‐proline (d ‐Pro‐l ‐Pro) dipeptide as the nucleating turn, and a thioether tether as a side‐chain linkage at a precisely designed position to stabilize the β‐hairpin. This method was used to modify the C‐terminal β‐hairpin moiety of the plant defensin, pv‐defensin, in order to obtain a stabilized peptide with enhanced anti‐Candida albicans activity (MIC 84–3.0 μm ), high serum stability (50 % remaining after 48 h) and low hemolysis (<10 % at 152 μm ). This modified peptide penetrated the C. albicans cell membrane within 5 min and showed high activity against clinically isolated antibiotic‐resistant C. albicans and Candida glabrata strains.     

Carbon assimilation and dissimilation during growth of Geotrichum candidum on amino acids and glucose

BACKGROUND: This work examines the metabolic behaviour of amino acids during Geotrichum candidum growth, in the presence of a primary carbon source like glucose. Amino acids were characterized based on their carbon assimilation and dissimilation by G. candidum, in the presence of glucose as the limiting substrate. RESULTS: The first group (Cys, His, Phe, Thr and Trp) was only used as nitrogen sources by G. candidum, with glucose being the carbon and energy source. Glucose repression was shown for the rest of the amino acids, since only after glucose depletion amino acids from the second group (Gly, Lys, Met, Val) were dissimilated for energy supply by oxidation into CO₂, while those from the third group (Ala, Arg, Asp, Glu, Leu, Pro and Ser) were assimilated as carbon sources (and additionally used as nitrogen sources), leading to a diauxic growth. CONCLUSION: This energy‐saving response was not previously shown for the second fungus involved in ripening of soft white cheese—P. camembertii—leading to simultaneous use of some amino acids and glucose as carbon and energy sources, and hence lower growth rates than those recorded during G. candidum growth. Copyright © 2007 Society of Chemical Industry     

Kinetics of amide formation on 1,4‐butanediol dimethacrylate crosslinked polystyrene resin: A comparison with PS‐DVB resin

An efficient crosslinked polymer support was synthesized by introducing 1,4‐butanediol dimethacrylate crosslinker to a polystyrene network using aqueous suspension polymerization technique. The support was functionalized with aminomethyl groups using chloromethyl phthalimide. The kinetics of amide bond formation on 1,4‐butanediol dimethacrylate crosslinked polystyrene (PS‐BDODMA) polymer were carried out using the attachment of Rink amide linker as a model reaction. The efficiency of the support was tested and compared with Merrifield resin by following different steps involved in the synthesis of a 14‐residue model peptide, Lys‐Ile‐Asn‐Thr‐Asn‐Ala‐Ser‐Trp‐His‐Ala‐Asn‐Arg‐Thr‐Ala‐NH_2, under the same synthetic conditions. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 88: 2897–2903, 2003     

Adsorption of certain amino acids onto crosslinked diethylaminoethyl dextran microbeads

The adsorption dynamism of certain amino acids [alanine (Ala), serine (Ser), lysine (Lys), proline (Pro), and aspartic acid (Asp)] on crosslinked diethylaminoethyl (DEAE) dextran microbeads was investigated with electronic spectroscopy in aqueous solutions. Among the selected amino acids, Ala‐Pro and Ser‐Lys showed similar adsorption dynamism, but Asp showed the strongest tendency for adsorption. The interaction dynamism of Asp with crosslinked DEAE dextran microbeads was studied to determine the mechanism of this adsorption. © 2005 Wiley Periodicals, Inc. J Appl Polym Sci 96: 1985–1991, 2005     

Synthesis of a Glycopeptide Vaccine Conjugate for Induction of Antibodies Recognizing O‐Mannosyl Glycopeptides

In spite of the clear importance of protein O‐mannosylation in brain glycobiology, tools are lacking for specific detection, enrichment, and identification of proteins containing these modifycations. We envisioned inducing antibodies that specifically recognize O‐mannose glycans on proteins and peptides. With this in mind, we prepared a glycopeptide vaccine construct containing the N‐acetyllactosamine‐extended mannose motif Galβ1‐4GlcNAcβ1‐2ManαThr, found as a common core structure on almost all mammalian O‐mannosyl glycoproteins identified. O‐mannose glycosylated amino acid building blocks and the corresponding glycopeptides were prepared by chemical synthesis and then conjugated to an immune carrier protein. After administration of the synthetic vaccine into rabbits, strong immune responses were obtained. Further evaluation by ELISA neutralization experiments and glycopeptide microarrays showed that the induced antibodies were highly specific to the glycopeptide antigen.     

Structural Rearrangement from Oligomer to Fibril Detected with FRET in a Designed Amphiphilic Peptide

β‐Sheet conformation is promoted in peptides with amphiphilic design, and stable β‐turn formation is favored with the unnatural amino acid d ‐Pro followed by a flexible residue such as Gly. A 19‐residue peptide (B3) was synthesized with alternating hydrophobic and hydrophilic residues connected by symmetrical d ‐Pro‐Gly and Gly‐d ‐Pro turns. B3 forms an oligomeric aggregate, rich in β‐sheet conformation, that reversibly transforms into an unordered structure on heating, as evidenced by its temperature‐dependent IR spectra. When a dansyl moiety was added to the N terminus of B3, the resulting peptide (B3D) can convert into a fibrillar structure after higher temperature incubation, as detected spectroscopically as well as by TEM. The fibrillization process involves an initial unfolding step monitored by the quenching of dansyl emission; in contrast, subsequent enhanced thioflavin T emission is seen on its binding to the fibril. A possible mechanism is proposed: B3D forms a low‐temperature oligomer, which is at least partially unfolded by heat and subsequently reassembles more slowly as a fibril.     

A Kinase‐Independent One‐Pot Multienzyme Cascade for an Expedient Synthesis of Guanosine 5′‐Diphospho‐d‐mannose

Biomimetic synthesis routes towards the important natural d ‐mannosyl donor guanosine 5′‐diphospho‐d ‐mannose (GDP‐Man) rely on kinase‐catalyzed nucleotide triphosphate (NTP)‐dependent phosphorylations of d ‐mannose (Man), to give d ‐mannose 6‐phosphate or α‐d ‐mannose 1‐phosphate (αMan 1‐P) as an intermediate product. A GDP‐Man synthesis not requiring the kinase/NTP system would be practical and cost‐effective. Here, we have developed a multienzyme cascade towards GDP‐Man, characterized in that αMan 1‐P was obtained by a diastereoselective phosphatase‐catalyzed phosphorylation of Man. α‐d ‐Glucose 1‐phosphate (αGlc 1‐P), prepared in situ through phosphorylase‐catalyzed conversion of sucrose in the presence of inorganic phosphate, was used as an expedient phosphoryl donor. The incipient αMan 1‐P and guanosine triphosphate (GTP) were converted into GDP‐Man by a highly manno compared to gluco selective nucleotidyltransferase. Pyrophosphatase was additionally required to hydrolyze the pyrophosphate released from the GTP, thus driving the reaction towards GDP‐Man. The enzymatic cascade was operated with the αMan 1‐P and the GDP‐Man formation decoupled from one another (sequential mode) or having all steps run concurrently (simultaneous mode). Detailed time course analysis revealed that kinetic pull due to the constant removal of the intermediate αMan 1‐P in simultaneous‐mode reactions was important to promote phosphorylation of Man from αGlc 1‐P in high efficiency, avoiding loss of sugar 1‐phosphates by hydrolysis. Under optimized conditions for the one‐pot transformation involving four enzymes, 100 mM (67 g L⁻¹) GDP‐Man was prepared from 140 mM sucrose and phosphate, using 400 mM Man as the phosphoryl acceptor. The product was recovered by anion‐exchange and size‐exclusion chromatography in ≥95% purity in about 50% yield (100 mg). These results demonstrate for the first time the practical use of a phosphorylase‐phosphatase combi‐catalyst as an alternative to the canonical kinase for the anomeric phosphorylation of the sugar substrate in nucleoside diphospho‐sugar synthesis. Phosphorylation from inorganic phosphate via the intermediate αGlc 1‐P rather than from NTP, particularly GTP, appears advantageous specifically in cases where the sugar acceptor is a bulk commodity that can be applied in suitable excess to the phosphatase reaction.

     

Expanding the Landscape of Amino Acid-Rich Antimicrobial Peptides: Definition,Deployment in Nature,Implications for Peptide Design and Therapeutic Potential

Unlike the α-helical and β-sheet antimicrobial peptides (AMPs), our knowledge on amino acid-rich AMPs is limited. This article conducts a systematic study of rich AMPs (>25%) from different life kingdoms based on the Antimicrobial Peptide Database (APD) using the program R. Of 3425 peptides, 724 rich AMPs were identified. Rich AMPs are more common in animals and bacteria than in plants. In different animal classes, a unique set of rich AMPs is deployed. While histidine, proline, and arginine-rich AMPs are abundant in mammals, alanine, glycine, and leucine-rich AMPs are common in amphibians. Ten amino acids (Ala, Cys, Gly, His, Ile, Lys, Leu, Pro, Arg, and Val) are frequently observed in rich AMPs, seven (Asp, Glu, Phe, Ser, Thr, Trp, and Tyr) are occasionally observed, and three (Met, Asn, and Gln) were not yet found. Leucine is much more frequent in forming rich AMPs than either valine or isoleucine. To date, no natural AMPs are simultaneously rich in leucine and lysine, while proline, tryptophan, and cysteine-rich peptides can simultaneously be rich in arginine. These findings can be utilized to guide peptide design. Since multiple candidates are potent against antibiotic-resistant bacteria, rich AMPs stand out as promising future antibiotics.     

Amino acids as carbon,energy and nitrogen sources for Penicillium camembertii

Three groups of amino acids were previously characterized on their ability to be assimilated as carbon source by Penicillium camembertii. In view of a deeper understanding of their metabolic behaviour, growth of P. camembertii on glucose, the limiting substrate, and an amino acid was examined in batch culture. Amino acids from the first group (Cys, His, Lys, Met, Trp and Val) are convenient nitrogen sources, but cannot be assimilated as carbon sources. However, they are also dissimilated, namely used for energy supply by oxidation into CO₂, during stationary phase, after glucose depletion, as shown for lysine; and the corresponding nitrogen was released as ammonium. Growth exhibited diauxic behaviour for the second group of amino acids (Arg, Leu), since they can be assimilated as carbon sources, in addition to their assimilation as nitrogen sources, but only after glucose depletion, as shown for arginine. A clear differentiation between the assimilated and the dissimilated carbon was demonstrated for the third group of amino acids (Ala, Asp, Glu, Gly, Pro, Ser, Thr); it was shown that the carbon from glutamic acid was assimilated, while the carbon from glucose was dissimilated. Copyright © 2006 Society of Chemical Industry