Block copolymers of amino acids. I. Synthesis and structure of copolymers of L-alanine or L-phenylalanine with D,L-lysine-d7 or D,L-lysine

Water-soluble block copolymers of the type (A)_m-(B)_n-(A)_p, where (A)_m,p was either poly(D ,L -lysine-α,β,β,γ,γ,δ,δ-d₇) or poly(D ,L -lysine) and (B)_n was either poly(L -alanine) or poly(L -phenylalanine), were synthesized for conformational studies by proton magnetic resonance spectroscopy. Analytical determination of the amount of the initiator fragment (n-hexylamine) at the C-terminus of the copolymers was used to obtain the number-average degrees of polymerization, DP _n, and thereby, together with the amino acid composition, to establish the covalent structures of the polymers. The values of DP _n were found to be much lower than those deduced from sedimentation equilibrium or form viscosity measurements. These deviations, which also are thought to have arisen in similar studies reported in the literature, are attributable to intermolecular aggregation; the relation of such aggregation to covalent structure (and its effect on the polymerization reaction) is discussed in terms of the conditions and mechanism of synthesis of block copolymers of amino acids.     

Renal transport of neutral amino acids. Cation-dependent uptake of L-alanine by luminal-membrane vesicles.

The characteristics of L-alanine transport in luminal-membrane vesicles isolated either from whole cortex or from pars convoluta or pars recta of rabbit proximal tubules were studied by a rapid filtration technique and by a spectrophotometric method. Uptake of L-alanine by vesicles from whole cortex was mediated by both Na+-dependent and Na+-independent, but electrogenic, processes. The nature, mechanism and tubular localization of the transport systems were studied by the use of vesicles derived from pars convoluta and pars recta. In vesicles from pars recta transport of L-alanine was strictly dependent on Na+ and occurred via a dual transport system, namely a high-affinity (half-saturation 0.14 mM) and a low-affinity system (half-saturation 9.6 mM). The cation-dependent but Na+-unspecific transport system for L-alanine was exclusively localized to the pars convoluta, which also contained an Na+-preferring system of intermediate affinity (half saturation 2.1 mM). A closer examination of the mechanism of transport of L-alanine in vesicles from pars convoluta revealed that an H+ gradient (extravesicular greater than intravesicular) can drive the transport of L-alanine into the vesicles both in the presence and in the absence of Na+. The physiological importance of various L-alanine transporters is briefly discussed.     

Enthalpy of helix-coil transitions from heats of solution. II. Poly-epsilon-carbobenzoxy-L-lysine and copolymers with L-phenylalanine

The heal of solution of Poly-ε-carbobenzoxy-L -lysine and of a series of its copolymers with phenylalanine was measured as a function of solvent composition. The enthalpy change of the helix-to-coil transition was estimated for the various cases. The previous findings that side chains do not greatly affect the transition enthalpy is confirmed also in cases having much larger differences in helix stability.     

Synthesis of polyhydroxy amino acids based on D- and L-alanine from D-glycero-D-gulo-heptono-1,4-lactone

2-Amino-2,3-dideoxy-D-manno-heptonic acid (7) has been synthesized from 2,5,6,7-tetra-O-acetyl-3-deoxy-D-gluco-heptono-1,4-lactone (1), which was readily prepared from D-glycero-D-gulo-heptono-1,4-lactone. O-Deacetylation of 1 followed by treatment with 13:1 (v/v) 2,2-dimethoxypropane/acetone in the presence of p-toluenesulfonic acid gave methyl 3-deoxy-4,5:6,7-di-O-isopropylidene-D-gluco-heptonate (3) as a crystalline product (80% yield). The free hydroxyl group (OH-2) of 3 was mesylated and substituted by azide to give the corresponding azide derivative 5. Hydrogenolysis and further hydrolysis of the ester function of 5 afforded alpha-amino acid 7 (43% overall yield from 1). Compound 7 is an analog of L-alanine having a polyhydroxy chain attached to C-3. The diastereoisomer of 7 at C-2, 2-amino-2,3-dideoxy-D-gluco-heptonic acid (12) was also prepared from 3, by a route that involved 2,3-dideoxy-2-iodo derivative 8 as a key intermediate.     

X-ray photoelectron spectroscopy of some selenium containing amino acids.

X-ray photoelectron spectra of some inorganic selenium compounds, Se-methionine, Se-cystine, Se-urea and selenodicysteine were recorded and compared with the XPS data obtained from the respective sulphur containing compounds. The oxidation state of selenium could be monitored by the observed chemical shifts of the Se(3p1/2),Se(3p3/2) and Se(3d3/2,5/2) levels. Though having a formal oxidation state near zero, the binding energy of the core electrons of Se in Se-methionine, Se-cystine and selenodicysteine was shifted by 0.4, 0.7 and 0.4 eV, respectively. This phenomenon was attributed to the rather distinct polarization of Se. The reversible oxidation of Se-cystine using H2O2 and NaBH4 could be successfully demonstrated by this XPS-technique.     

Design and engineering of metalloproteins containing unnatural amino acids or non-native metal-containing cofactors

An emerging branch of metalloprotein design and engineering is on the horizon, where unnatural amino acids or non-native metal-containing cofactors are employed in the design and engineering process. These endeavors have been shown to be quite effective in elucidating the precise roles of key residues in protein structures and functions, in providing guiding principles on protein design, in fine-tuning the protein properties to an unprecedented level, and in expanding the repertoire of protein functionalities, and thus its range of applications.     

Polymerization of amino acids containing nucleotide bases

Summary We have prepared the nucleoamino acids 1-(3-amino, 3-carboxypropyl)uracil (3) and 9-(3-amino, 3-carboxypropyl)adenine (4) as (l)-enantiomers and as racemic mixtures. When3 or4 is suspended in water and treated with N,N-carbonyldiimidazole, peptides are formed in good yield. The products formed from the (l)-enantiomers are hydrolyzed to the monomeric amino acids by pronase.Attempts to improve the efficiency of these oligomerizations by including a polyuridylate template in the reaction mixture were not successful. Similarly, oligomers derived from the (l)-enantiomer of3 did not act as templates to facilitate the oligomerization of4.     

Thia-analogues of amino acids synthesis of peptide derivatives containing 3-thia-analogues of amino acids

Summary N-Protected dipeptides containing L-3-thia-analogues of phenylalanine, p-nitro-phenylalanine, lysine and leucine respectively were prepared applying an enantioselective enzymatic reaction step. Racemic synthetic intermediates of the type acyl-NH-CH(R¹)-CO-D,L-NH-CH(S-R²)-COOBzl were selectively deprotected at the C-terminus by enzymatic hydrolysis using thermitase or trypsin.Abbreviations Ac acetyl - AcOEt ethyl acetate - AcOH acetic acid - Boc tert.-butyloxycarbonyl - Bz benzoyl - Bzl benzyl - DMF dimethyl-formamide - EtOH ethanol - THF tetrahydrofuran - Z benzyloxycarbonyl Dedicated to Prof. D. Cavallini at the occasion of his 75th birthday.     

Shock synthesis of amino acids II

The synthesis of amino acids behind shock waves in methane, ethane, ammonia, and water vapor was further investigated. Aldehydes and HCN and formed separately in the non-homogeneous gas during the high temperature period and recombine with ammonia during the thermal quench period, to form -amino nitriles. The -amino nitriles are either hydrolyzed by excess water vapor during the quench period or emerge as such after the reaction is completed. A combined gas chromatograph-mass spectrometer analysis of the reaction product showed identical amounts of D and L amino acids, thus confirming the absence of contaminants. Thunder shock waves were shown to be a suitable source of energy for the production of amino acids.     

Site-specific PEGylation of proteins containing unnatural amino acids

Here, we report a generally applicable PEGylation methodology based on the site-specific incorporation of para-azidophenylalanine into proteins in yeast. The azido group was used in a mild [3+2] cycloaddition reaction with an alkyne derivatized PEG reagent to afford selectively PEGylated protein. This strategy should be useful for the generation of selectively PEGylated proteins for therapeutic applications.     

ESR an optical absorption studies on the copper(II) interaction with small peptides containing aromatic amino acids.

The interaction of Cu(II) with di- and tripeptides each containing phenylalanine, tryptophan or histidine in the amino acid chain has been investigated by means of electron spin resonance (ESR) and optical absorption spectroscopy. Cu(II) complexes of dipeptides and tripeptides exhibit different magnetic and optical parameters. Dipeptide complexes have larger gparallel-values and smaller A parallel values than tripeptide complexes. When compared to dipeptide complexes, the d-d band of the central metal ion is blue shifted for tripeptide complexes. There are no significant difference in the behavior of Cu(II) peptide complexes containing phenylalanine or tryptophan. Complexes of histidine containing peptides, however, show modified spectra caused by the participation of the imidazole nitrogen in the coordination to Cu(II). The imidazole nitrogen seems to coordinate in-plane with other coordinating atoms or in an axial position depending on the kind of peptide.     

Solution and solid state study of copper(II) ternary complexes containing amino acids of interest for brain biochemistry - 1: Aspartic or glutamic acids with methionine or cysteine

Four new ternary complexes of Cu(II) containing aspartic or glutamic acids and methionine or cysteine as ligands were studied both in solution and in solid state. The solution study was performed using potentiometry, UV-Vis spectroscopy, electron paramagnetic resonance (EPR) spectroscopy, and electrochemistry. The stability constants of the ternary complexes were determined by potentiometry. The order of the values for the stability constants was CuGluMet (complex 4) < CuAspMet (complex 2) < CuGluCys (complex 3) < CuAspCys (complex 1), with complex 1 ≈ complex 3 ? complex 2 ≈ complex 4. This behavior was confirmed by the electronic spectra, EPR parameters and cyclic voltammetry. The compounds were synthesized in the solid state and characterized by thermogravimetry as well as vibrational and EPR spectroscopy. The results found in solid state are in agreement with the data found in solution studies where methionine, aspartic and glutamic acids coordinate to the copper(II) ion using the nitrogen atom of the amino group and one oxygen atom of the carboxylate group and cysteine coordinates through the nitrogen and sulfur atoms. The EPR data suggest that the complexes are square planar and that the complexes with cysteine as one of the ligands have some distortion.     

Inhibition of beta-amyloid toxicity by short peptides containing N-methyl amino acids.

Single N-methyl amino acid-containing peptides related to the central hydrophobic region beta16-20 (Lys-Leu-Val-Phe-Phe) of the beta-amyloid protein are able to reduce the cytotoxicity of natural beta1-42 in PC12 cell cultures. N-methyl phenylalanine analogs yield statistically significant increments in cell viability (Student's t-test < 0.01%) and are nontoxic in the same assay. These promising results indicate that these peptide molecules could be a starting point for the development of potential therapeutic compounds for the treatment of Alzheimer's disease.