Insect Trypsin Modulating Oostatic Factor (Neb-TMOF) and Its Analogs: Preliminary Structure/Biological Function Relationship Studies

Summary Neb-TMOF, the trypsin modulating oostatic factor of gray fleshflyNeobellieria bullata, is a hexapeptide with the following sequence: H-Asn-Pro-Thr-Asn-Leu-His-OH. It has been isolated from vitellogenic ovaries in 1994. TMOF, the newly discovered insect peptide, inhibits trypsin biosynthesis in the gut, lowers yolk polypeptide concentration in the hemolymph and strongly inhibits ecdysone biosynthesis by larval ring glands. It is interesting that this short non-protected peptide contains in its molecule two Asn residues at positions 1 and 4 and His at its C-terminus. To obtain information about the role of the His-6 and Asn-4 residues we synthesised two series of Neb-TMOF analogs, modified: (1) in position 6 byd-His (I), His(Bzl) (II) and Phe(p-X) derivatives, where X=NH₂ (III), NO₂ (IV), OEt (V) and OH (VI) and (2) in position 4 by such amino acid residues as Ser (VII), Thr (VIII), Gly (IX), Asp (X), Glu (XI) andd-Asn (XII). The influence of these peptides on trypsin biosynthesis inN. bullata was determinedin vivo. In preliminary investigations, we found that Neb-TMOF, [Phe(NH₂)⁶], and [Phe(NO₂)⁶]-Neb-TMOF inhibited trypsin biosynthesis, whereas [d-His)⁶]- and [d-His(Bzl)⁶]-Neb-TMOF were inactive. In further biological studies performedin vitro on heart ofTenebrio molitor were found that-TMOF and [Phe(p-NH₂)⁶]-Neb-TMOF showed weak cardioexcitatory activity, about 30% of the cardioexcitatory activity of proctolin, an insect neuromodulating peptide.     

Thiophosphate analogs of ADP and ATP as substrates in partial reactions of energy conversion in chloroplasts

Thiophosphate analogs of ADP and ATP have been employed in partial reactions of photosynthetic energy conversion in chloroplasts. Substitution of oxygen by sulfur at the α-phosphate yields a pair of diastereomers (ADPαS, ATPαS, A and B forms). Two diastereomeric compounds are also obtained by replacement of oxygen by sulfur in the β-phosphate group of ATP (ATPβS, A and B form) (Eckstein, F. and Goody, R.S. (1976) Biochemistry 15, 1685–1691).The A form of ADPαS is phosphorylated by chloroplasts with a K_m comparable to that of ADP but with a lower V. The B form of ADPαS as well as ADPβS is not a substrate in photophosphorylation and only weakly competes with ADP.The A forms of ADPαS and ATPαS strongly compete with ADP for the tight nucleotide binding site of CF₁ in the light-induced exchange reaction, whereas the B forms display a much smaller competitive effect. The efficiencies of ADPβS and the A isomer of ATPβS are intermediate, and the B form of ATPβS is a weaker competitor.The A forms of ATPαS and ATPβS are hydrolyzed by light-triggered ATPase, whereas the B forms are not. The efficiency of the A isomer of ATPαS is comparable to that of normal ATP, and the A form of ATPβS is cleaved at a lower rate. In trypsin-activated Ca²⁺-dependent ATPase the A form of ATPαS is the only thiophosphate analog to be hydrolyzed.The results indicate a stereospecific interaction of ADP and ATP at the catalytic sites as well as the tight nucleotide binding site of coupling ATPase of chloroplasts.     

Mast cell degranulating (MCD) peptide analogs with reduced ring structure

Mast cell degranulating (MCD) peptide, a component of bee venom, is a 22 amino acid peptide with two disulfide bridges. In this first structure-activity study of MCD peptide, three analogs were synthesized and tested: two analogs shortened by omitting sequences 6–10 and 8–13, respectively, and one analog lacking the disulfide bridge between cysteine residues 5 and 19. These analogs were synthesized by solid-phase methods and were compared to MCD peptide in two assays for inflammation: histamine release from mast cells and superoxide anion release from neutrophils. All three analogs produced histamine release, although with only about one fifth of the activity of MCD peptide. Superoxide anion-releasing activity, however, did not parallel histamine release. MCD peptide did not release superoxide anion, while the 6–10 and 8–13 deletion analogs were strong and weak stimulants, respectively, of this anion. CD spectra showed that the secondary structures of the three analogs were very similar to that of MCD peptide, so that a change in secondary structure cannot completely explain the changes in releasing activities. Charge differences between the two deletion analogs and MCD peptide may explain some of the differences in activity. This is the first demonstration that the various activities of MCD peptide can be separated, and provides a lead through which the purported antiinflammatory activity of MCD peptide may possibly be explored in the future.     

Direct enantioselective separation of some propranolol analogs by HPLC on normal and reversed cellulose chiral stationary phases

The enantiomeric separation of several racemic aryloxyaminopropan-2-ol derivatives related to propranolol on normal and reversed phase of cellulose tris (3,5-dimethylphenylcarbamate) chiral stationary phases known as Chiralcel OD and Chiralcel OD-R were studied. It was observed that the chiral separation depends on the substitution pattern of the aryl group, i.e., 1-naphthyl, 2-naphthyl, and phenyl group and polarity on the basic nitrogen in the side chain. In both normal and reversed phase modes the (+)-R-enantiomer eluted first in all of the analogs resolved. It can be concluded that: (1) substituents on the side chain did affect the interaction of the enantiomers with the polar carbamate moiety in the CSP; and (2) the dipole-dipole stacking between the π-donor 3,5-dimethylphenyl carbamate group pending from the glucose rings of the CSP and π-acceptor aryl group of the analyte is crucial for the efficient chiral discrimination. The chiral recognition mechanism(s) between these analogs and the chiral stationary phases are proposed. © 1996 Wiley-Liss, Inc.     

Structure-activity analyses reveal the existence of two separate groups of active octadecanoids in elicitation of the tendril-coiling response of Bryonia dioica Jacq.

The Bryonia dioica tendril-coiling assay provides a rapid, sensitive and selective bioassay for jasmonates. Using this assay, a large number of jasmonate and coronatine analogs were analyzed for their biological activities. In a systematic study, C-3 analogs, C-2 analogs, C-1 homologs and -analogs, C-1(1′) analogs of jasmonic acid, as well as analogs of coronatine altered in both the amino acid and the coronafacic acid moiety, were compared. The results demonstrated at least two structurally non-overlapping centers of biological activity, one centered around the structure of jasmonic acid allowing only minor C-1(1′) modifications and a second center around the structure of 12-oxophytodienoic acid and having different structural requirements for activity, thus allowing quite different structural modifications. The C18-group of the jasmonates [12-oxophytodienoic acid and 3-oxo-2(2′ (Z)-pentenyl)-cyclopentane-1-octanoic acid], for which coronatine is a structural mimic, was the much more potent inducer of tendril coiling, when applied externally. The levels of jasmonic acid and 3-oxo-2(2′(Z)-pentenyl)-cyclopentane-1-octanoic acid in mechanically stimulated tendrils remained very low and did not change detectably, while the level of 12-oxophytodienoic acid had earlier been shown to change drastically and transiently during the onset and progression of the coiling reaction. Thus, 12-oxophytodienoic acid, rather than jasmonic acid or 3-oxo-2(2′ (Z)-pentenyl)-cyclopentane-1-octanoic acid, has to be considered as an endogenous signal transducer in B. dioica mechanotransduction. Received: 19 June 1998 / Accepted: 14 September 1998     

抗病毒核苷酸类似物磷酸化修饰研究进展

核苷(酸)类似物是一类抗病毒前药,其进入人体细胞后经过逐步磷酸化生成核苷三磷酸类似物发挥抗代谢药作用,主要通过抑制病毒复制和促进侵染细胞凋亡,达到疾病治疗效果.其中,核苷类似物在细胞内经激酶活化的代谢转化过程通常是不充分的,导致最后生成的核苷三磷酸类似物浓度较低,降低了作用效果.因此,通过直接制备核苷酸类似物作为抗病毒...     

Binding of the new morphiceptin analogs to human MCF-7 breast cancer cells and their effect on growth

In the present study, we reported on the synthesis of two new mu-opioid peptide analogs, [D-1-Nal3]morphiceptin and [D-1-Nal4]-morphiceptin [1-Nal=3-(1-naphthyl)-alanine] which expressed receptor binding affinities at least at the level of the primary opioid ligands. The new analogs also labeled mu-opioid receptors on the cells of human breast cancer MCF-7 cell line with affinity much higher than that of endomorphins and morphiceptin, the well-known mu-selective opioid peptides. However, none of the tested peptides significantly decreased cell proliferation of MCF-7 cells.     

Orientation and Mobility of Actin in Different Intermediate States of the ATP Hydrolysis Cycle

Using polarization fluorimetry, we have investigated conformational changes of FITC-phalloidin-labeled F-actin in ghost muscle fibers. These changes were induced by myosin subfragment-1 (S1) in the absence and presence of MgADP, MgAMP-PNP, MgATPgammaS, or MgATP. Modeling of various intermediate states was accompanied by discrete changes in actomyosin orientation and mobility of fluorescent dye dipoles. This suggests multistep changes of orientation and mobility of actin monomers during the ATPase cycle. The most pronounced differences in orientation (~4 degrees ) and in mobility (~43%) of actin were found between the actomyosin states induced by MgADP and MgATP.     

New conformationally restricted analog of the immunosuppressory mini-domain of HLA-DQ and its biological properties

Our previous studies revealed that the nonapeptide fragment of HLA-DQ located in the beta 164-172 loop of the Thr-Pro-Gln-Arg-Gly-Asp-Val-Tyr-Thr sequence suppresses the immune humoral and cellular responses [30]. Based on the crystal structure of HLA-class II molecules we designed and synthesized a cyclic analog with restricted conformation, cyclo(Suc-Thr-Pro-Gln-Arg-Gly-Asp-Val-Lys)-Thr-OH (Suc = succinyl) by reacting a Lys side chain with a succinylated N-terminus. The cyclization product more potently suppresses the cellular immune response than its linear counterparts and is efficiently cleaved by trypsin. The results indicate that the beta 164-172 loop may serve as a functional epitope on the HLA class II surface for intermolecular binding.