Tissue-type transglutaminase is irreversibly inactivated during heat treatment. The rate of inactivation is low at pH 7.5; it increases slightly at acid pH (6.1) but much more at alkaline pH (9.0±9.5), suggesting that specific effects take place in the alkaline range, possibly in relation to decreased stability of the transition-state intermediate as pH is raised above 9.0. Differential scanning calorimetry experiments indicate that thermal unfolding of the protein occurs with two separate transitions, involving independent regions of the enzyme. They are assigned to domains 1 and 2 and domains 3 and 4, respectively, by a combination of calorimetric and spectroscopic techniques. When considering the effects of pH, we noted that transglutaminase was unfolded via different pathways at the different pH values considered. At acid pH, the whole structure of the protein was lost irreversibly, with massive aggregation. At neutral and, even more so, at alkaline pH, aggregation was absent (or very limited at high protein concentration) and the loss of secondary structure was dependent on the ionization state of crucial lysine residues. Unfolding at pH 9.5 apparently chiefly involved the N-terminal region, as testified by changes in protein intrinsic fluorescence. In addition, the C-terminal region was destabilized at each pH value tested during thermal unfolding, as shown by digestion with V8 proteinase, which is inactive on the native protein. Evidence was obtained that the N-terminal and C-terminal regions interact with each other in determining the structure of the native protein.

The effect of pH and temperature on the association equilibrium constant (Ka) for the binding of the trypsin inhibitor from white mustard (Sinapis alba L.) seeds (MTI) to bovine beta-trypsin (EC 3.4.21.4) has been investigated. On lowering the pH from 9 to 3, values of Ka for MTI binding to bovine beta-trypsin decrease thus reflecting the acid-pK and -midpoint shifts, upon inhibitor association, of two independent ionizable groups, and of a three-proton transition, respectively. At pH 8.0, values of thermodynamic parameters for MTI binding to bovine beta-trypsin are: Ka = 4.5 X 10(8)M-1, delta G0 = -11.6 kcal/mol, and delta S0 = +53 entropy units (all at 21 degrees C); and delta H0 = +4.1 kcal/mol (temperature independent between 5 degrees C and 45 degrees C). Binding properties of MTI to bovine beta-trypsin have been analyzed in parallel with those concerning macromolecular inhibitor association to serine (pro)enzymes.

The synthesis and a conformational study of a number of homologues of the well known antibiotic, phytotoxic leucinostatin A are reported. The circular dichroism of all the compoundsare discussed. Someconclusions on the SARof these compoundsare drawn. The influence of the a-helical conformation and/or the increased lipophile character on their interesting biological activities is emphasized.

Among the compounds discussed for anti-microbial and anti-fungal use allicin (allylthiosulfinate, diallyl disulfide-S-monoxide), an active ingredient of garlic, has attracted considerable attention. The objective of this study is to determine the antifungal activity of a local garlic ecotype (Voghiera) extracts against different pathogens. Primary screening was carried out by the agar plates technique using ethanol garlic extract at four final concentrations against the following organisms: Alternaria alternata, Aspergillus spp., Colletotrichum acutatum, Didymella bryoniae, Fusarium culmorum, Fusarium avenaceum, Fusarium gramineareum, Gliocladium roseum 47, Pythium splendens, Rhizoctonia solani, Sclerotium rolfsii, Stemphylium vesicarium, Trichoderma longibranchiatum, and Botrytis cinerea. Secondary screening was carried out using a lyophilized and a spray-dried preparation at different concentrations against the organisms selected for the high inhibition garlic effect in the primary screening and compared with the commercial fungicides mancozeb and iprodione. The best results were observed for the spray-dried garlic compound that showed a good fungicidal activity at the concentration of 1.5 g/10 mL while lyophilized garlic at the same concentration exhibited less inhibition activity against the four fungi analyzed in the second screening.

The formylpeptides formyl-methionyl-N-methylleucyl-phenylalanine methyl ester [for-Met-(NMe)Leu-Phe-OMe] 1, formyl-methionyl-2-aminotetralin-2-carboxyl-phenylalanine methyl ester [for-Met-Atc-Phe-OMe] 2, formyl-methionyl-1,2,3,4-tetrahydroisoquinoline-3-carboxyl-phenylalanine methyl ester [for-Met-Tic-Phe-OMe] 3 and formyl-methionyl-2-aminoxy-4-methylvaleryl-phenylalanine methyl ester [for-Met-OLeu-Phe-OMe] 4 were synthesized in order to investigate the role of the amide bond at position 2 on biological activities on human neutrophils. Only analogue 2, which keeps the NH group at position 2, was found to retain activity though sterically encumbered.