Ivailo Ivanov

Enzymic modification of feather keratin hydrolysates with lysine aimed at increasing the biological value

The lysine content of feather protein hydrolysate was modified via activation with alkaline proteinase and trypsin at pH 6·2 and 8·3, respectively. Lysine was used as the hydrochloride and the diacetyl derivative in the first case and as the methyl ester in the trypsin modification. The results indicate a lysine content of 26–35 g kg-1 and 59–64 g kg-1 after alkaline proteinase activation and 52–65 g kg-1 after trypsin activation. Considerable changes in the molecular mass profile of peptides were observed. Thus, enzyme modification is a way to raise the nutritive value of feather protein hydrolysate by increasing its lysine content (the main limiting essential amino acid).

Enzymic Synthesis Design and Enzymic Synthesis of Aspartame

Abstract An enzymic synthesis of aspartame (H-Asp-Phe-OMe) has been designed and realized based on the structure-activity study of thermolysin and penicillin amidase hydrolysis of its p-substituted phenylacetyl derivatives. These compounds meet the structural and energetic requirements of two enzymic binding sites The peptide sweetener has been prepared by thermolysin - catalyzed condensation of the p-substituted phenylacetyl-Asp-OH and H-Phe-OMe followed by penicillin amidase - catalyzed deprotection of the resulted aspartame precursors.

Ortho—Nitrophenol Removal in two Types Activated Sludge: The Role of Microbiological and Enzymological Adaptation

ABSTRACTThe relation between the structural and functional changes in two types anaerobic activated sludge—ASt (activated sludge trivial) and ASg (activated sludge granular) from waste water treatment process with high organic load and working under presence of hazardous pollutants (oNP—ortho-nitrophenol) has been investigated. The adaptive response on the different levels—microbiological structure, effectiveness of oNP-removal, oxygenase and succinate dehydrogenase activities towards the increasing oNP-concentration has been assessed. The obtained results demonstrate that for the complete oNP-biodegradation to harmless products—the aerobic finishing treating process is applicable and needed. In these conditions the amount of microorganisms from genus Pseudomonas and oNP-degrading bacteria, an efficiency oNP-biodegradation and oxygenase activities were increased considerably.

Thermodynamics of enzymic peptide synthesis in biphasic aqueous-organic systems : Coupling of the chemical and phase equilibria

Chemical and phase equilibria in enzymic peptide synthesis in a biphasic aqueous–organic system are coupled, the mass transfer providing the “driving force” for the synthetic reaction. The overall synthetic equilibrium is characterized by an equilibrium constant, independent of pH, phase volume ratio and reactants partition coefficients. A linear free-energy relationship is observed for this equilibrium constant and the extraction equilibrium constant.

Xenobiotic Biotransformation Potential of Pseudomonas Rhodesiae KCM-R5 and Bacillus Subtilis KCM-RG5, Tolerant to Heavy Metals and Phenol Derivatives

ABSTRACT Two environmental bacterial isolates KCM-R5 and KCM-RG5 were selected from xenobiotic-polluted environment. KCM-R5 was identified as Pseudomonas rhodesiae and KCM-RG5 as Bacillus subtilis. KCM-R5 demonstrated tolerance to heavy metals and KCM-RG5—to heavy metals and phenol derivatives. Both strains were studied for xenobiotic biotransformation in order to contribute towards bioremediation of polluted environments. Pseudomonas rhodesiae KCM-R5 and Bacillus subtilis. KCM-RG5 possess unusual ability to utilize ortho-nitrophenol (o-NP) and 2,4- dichlorophenoxyacetic acid (2,4-D). o-NP and 2,4-D were added at concentration 30 mg/l. The possible indictive/inhibiting effect of Pb cations (40mg/l) was also studied. Pseudomonas rhodesiae KCM-R5 removed 86% of o-NP and below 1% of 2,4-D. Bacillus subtilis KCM-RG5 eliminated 83% of o-NP and under 1% of 2,4-D. Biotransformation effectiveness of o-NP reached 95–100% in contrast to 2,4-D where the effectiveness was just 15–20%. Cell morphological changes were registered during the biotransformation processes. The obtained results could contribute to manage bioremediation processes in polluted with heavy metals and phenol derivatives environments.

Thermodynamics of enzymic synthesis of solid-phase peptides

Abstract The Gibbs free energy changes of the individual and net synthetic equilibrium of solid-phase tert-butyloxycarbonyl-Phe-Gly p-substituted anilides are calculated from the HPLC analysis data for the equilibrium concentrations. A linear free energy relationship is observed for the net synthetic equilibrium and precipitation equilibrium, suggesting that the latter provides the driving force for the chymotryptic synthesis of these insoluble peptides. This conclusion is strongly supported by a linear correlation between the synthetic yield and the square root of the product solubility.

Structural/Functional Changes in Activated Sludge in PCP- and ONP-Biodegradation Technologies

ABSTRACT Different detoxication technologies (aerobic, anaerobic, two stage, hybrid) using oNP, PCP, DCP, TCP like model xenobiotics, have been accomplished in the last 8 years. The results about the details of the technologies have been described previously. In this paper the comparison between the efficiency of the different technologies and structural/functional changes of AS (activated sludge) has been made. The paper is illustrated by means of microscopic and transmission microscopic photographs. The results showed well expressed connection between the efficiency of xenobiotic elimination, type of technology, initial concentration of the particular xenobiotic and adaptive structure of AS. At aerobic shorter adaptation genus Pseudomonas played more important role in the detoxication. At time of the longer anaerobic and anoxic processes genus Acinetobacter assumed the govern role in the structure of xenobiotic-degrading bacterial complex. The deep knowledge of the adaptive structure of the different detoxication technologies presents the valuable information for quick and intelligent management of the detoxication bioremediation technologies.

Pentachlorophenol Biodegradation in Anaerobic and Aerobic Continuous Flow Processes

ABSTRACT Pentachlorophenol (PCP) biodegradation was studied comparatively in anaerobic and aerobic conditions. Two laboratory scale systems of completely mixed continuos flow bioreactor and secondary clarifier were used. Simultaneous treatment of municipal and industrial wastewater with high concentration of PCP (25 mg/l) was investigated. The results show that the biodegradation of trivial organic and PCP in aerobic conditions was better than in anaerobic conditions: the removal efficiencies were 83,3%(COD) and 55%(PCP) in the aerobic system against 41,3% (COD) and 47%(PCP) in the anaerobic system. The mean values of the effluent concentrations were 337 mg/l (COD) and 11 mg/l (PCP) for the aerobic experiment against 1263 mg/l (COD) and 12,8 mg/l (PCP) for the anaerobic experiment. Although the removal efficiencies of PCP and COD were relatively good for the investigated extremely high influent PCP concentration, both schemes did not meet the standards for the effluent concentration. Therefore the investigated bioreactors could be used only as a first stage in a system of bioreactors for completely PCP removal.

Antibody-Catalysed Peptide Nitroanilide Hydrolysis Using a new type of Hapten

Two monoclonal antibodies obtained after immnunization of BALB/c mice with Gly-Gly-Arg-semicarbazone coupled to BSA were found to catalyse the hydrolysis of Z-Gly-Gly-Arg-pNA. To explain this it was assumed (1) that the semicarbazone was converted to the corresponding hydrated tripeptide aldehyde by the chemical entities encountered in vivo during immunization (2) that it structurally mimics the transition-state(s) of amide hydrolysis and (3) that the presence of a charge in the hapten may not be crucial. The presence of the two lone pair orbitals on the O atoms from the OH groups could give rise to an “oxyanion hole”-like structure in the antibody combining site as in the serine proteinases. Both antibodies show high substrate discrimination. They do not catalyse the hydrolysis of Ac-Leu-Leu-Arg-pNA, while for trypsin the ratio between the second order rate constants of these substrates is only 2.3. On the other hand, however, the difference in the specificity constant for 2C9 and trypsin for Z-Gly-Gly-A...

Semispecific Ester Substrates of Trypsin as Acyl Donors in Kinetically Controlled Peptide Synthesis

Esters of benzoyl-Phe with different size and structure of the leaving group have been synthesized and their acyl-donor properties studied in tryptic kinetically controlled peptide synthesis of model peptides. The results obtained show that: (1) part of the leaving group of these substrates is involved in S12–P12interactions in the extended active site of the enzyme; (2) both hydrophobic and H-bond interactions contribute to the S12–P12-interactions; (3) the lack of secondary hydrolysis provides higher synthetic yields as compared to those when using specific trypsin substrates in kinetically controlled peptide synthesis.