X-ray crystal structure of papain complexed with cathepsin B-specific covalent-type inhibitor: substrate specificity and inhibitory activity.

The Ile-Pro sequence of CA074, potent covalent-type inhibitor, is necessary to exhibit the specificity for cathepsin B, but not for papain. In order to elucidate how its sequence binds to papain and why such binding does not exhibit the specificity for papain at the atomic level, two CA074-related compounds, 1 (N-(L-3-carboxyloxirane-2-carbonyl)-L-isoleucyl-L-proline) and 2 (N-(L-3-carboxyloxirane-2-carbonyl)-L-isoleucyl-diethylamide), were designed and their structure--inhibitory activity relationship was investigated by the X-ray crystal analyses of the complexes with papain. The Ile-Pro moiety of 1 was located at the S2 and S3 subsites consisting of Val-133, Val-157, and Asp-158 and of Tyr-61, Gly-66, and Tyr-67 residues of papain, respectively, which is in contrast with the binding of CA074 to S'n (n = 1 approximately 2) subsites in the complex with cathepsin B. Although 2 in the complex with papain showed the similar binding pattern to 1, its inhibitory activity was about two-fold higher than of 1, suggesting the importance of tight S3-P3 hydrophobic interaction for the activity. The difference of the substrate specificity between papain and cathepsin B has also been discussed based on the X-ray results of the present and cathepsin B-inhibitor complexes.

Long-term histological evaluation of hydroxyapatite ceramics in humans.

We have investigated the interface between bone and hydroxyapatite (HA) chronically implanted in man. By light microscopy, HA appeared to bind directly to bone without intervening fibrous tissue. By transmission electron microscopy, two patterns were noted: (1) HA either bound directly to bone; or (2) electron-dense material intervened between HA and bone. The orientation of bone collagen fibres likewise showed two patterns: (1) collagen fibres were oriented parallel to the HA; or (2) the fibres were aligned perpendicularly. We have observed similar binding properties of HA to the jaw bone of humans in vivo and in vitro.

Studies on the ionophorous antibiotics. XXVI. The assignments of the 13C-NMR spectra of lonomycin A and mutalomycin.

All resonances observed in the 13C-nmr spectra of the polyether antibiotics lonomycin A and mutalomycin have been assigned by the aid of biosynthetic method, selective proton decoupling as well as comparison with structurally related compounds.

NG-011 and NG-012, novel potentiators of nerve growth factor. II. The structure determination of NG-011 and NG-012.

NG-011 and NG-012, novel potentiators of nerve growth factor (NGF) were isolated from the culture broth of Penicillium verruculosum F-4542. The structures of NG-011 and NG-012 were elucidated by their spectral analysis and degradation experiments as shown in Fig. 1.

Microbial glycosylation of macrolide antibiotics by Streptomyces hygroscopicus ATCC 31080 and distribution of a macrolide glycosyl transferase in several Streptomyces strains.

In the course of our microbial transformation study on erythromycin derivatives, Streptomyces hygroscopicus ATCC 31080, which produces a polyether antibiotic carriomycin, was found to transform erythromycin derivatives to their inactivated derivatives. The structures of inactivated derivatives prepared by enzyme reaction using the cell extract, UDP-glucose (or UDP-galactose) and Mg2+ (or Mn2+) were elucidated on the basis of analysis of thei spectral data to be the compounds glycosylated at C-2' of a desosamine moiety, indicating that the enzyme is a macrolide glycosyl transferase (MGT). The MGT activity of cell extract from S. antibioticus ATCC 11891, a producing organism of oleandomycin, could be distinguished from that of ATCC 31080, based on the ability to glycosylate tylosin. We examined 32 actinomycete strains producing such polyketides as macrolide and polyether antibiotics, and found that 15 strains of Streptomyces have macrolide glycosyl transferase activity. It suggests that the MGTs have been distributed among at least polyketide producing Streptomyces strains.

Structures of new 18-membered macrolides FD-891 and FD-892.

Structures of FD-891 and FD-892 were determined by extensive NMR spectral analysis as shown in Fig. 1. They belong to such 18-membered macrolides as concanamycins and virustomycin.

NG-011 and NG-012, novel potentiators of nerve growth factor. I. Taxonomy, isolation, and physico-chemical and biological properties.

NG-011 and NG-012, novel potentiators of nerve growth factor (NGF), were isolated from the culture broth of Penicillium verruculosum F-4542, together with 3,4-dihydro-6,8-dihydroxy-3-methylisocoumarin. They potentiated the neurite outgrowth induced by NGF in rat pheochromocytoma cell line (PC12).

Microbial transformation of 6-O-methylerythromycin derivatives.

Mucor circinelloides f. griseo-cyanus IFO 4563 was found to convert 6-O-methylerythromycin A (TE-031, A-56268) to (14R)-14-hydroxy-6-O-methylerythromycin A [14R)-14-hydroxy TE-031). The TLC and spectral data of the conversion product were perfectly identical with those of an active major metabolite of TE-031 in humans (M-5). A related antibiotic, 6-O-methylerythromycin B (TB-010), was able to be similarly transformed to its C-14 hydroxy analogue [14R)-14-hydroxy-6-O-methylerythromycin B, (14R)-14-hydroxy TB-010). The MICs of (14R)-14-hydroxy-6-O-methylerythromycin B against some Gram-positive bacteria were almost equal to those of 6-O-methylerythromycin B. It is suggested that the hydroxylation at C-14 of 6-O-methylerythromycins A and B scarcely reduces their in vitro activity.

Stachybotrin C and parvisporin, novel neuritogenic compounds. II. Structure determination.

The structures of stachybotrin C and parvisporin have been determined by spectroscopic analyses and chemical derivatization. Stachybotrin C contains a unique pyrano-isoindolinone ring system, while parvisporin has a hydroxyl farnesyl phenol structure.

Isolation and characterization of a new pyrano[4',3':6,7]naphtho[1,2-b]xanthene antibiotic FD-594.

During our screening of microbial metabolites for effective drugs against tumor cell lines, we discovered a new pyrano[4',3':6,7]naphtho[1,2-b]xanthene derivative, FD-594 from the fermentation broth of Streptomyces sp. TA-0256. FD-594 shows moderate activity against tumor cell lines, comparative to that of adriamycin, as well as antibacterial activity against some Gram-positive bacteria.

Structure and biosynthesis of FD-594; a new antitumor antibiotic.

The structure of a novel antitumor antibiotics FD-594 (1), produced by Streptomyces sp. TA-0256, was determined to have a glycosylated pyrano[4',3':6,7]naphtho[1,2-b]-xanthene skeleton by means of spectral data. The biosynthetic studies of the chromophore of 1 was also carried out by feeding experiments with [1-13C]-, [2-13C]-, and [1,2-13C2]sodium acetate. The labeling pattern was determined by 13C NMR including 2D INADEQUATE experiments, which allowed us to elucidate that the chromophore of 1 is derived from 14 acetate, followed by the loss of one carbon atom.

Isolation and characterization of new 18-membered macrolides FD-891 and FD-892.

New 18-membered macrolides FD-891 and FD-892 were discovered from the fermentation broth of Streptomyces graminofaciens A-8890 isolated from a soil sample collected at Yamanashi prefecture Japan. They induce morphological changes of HL-60 cells at low concentration below IC50s and have cytocidal activity against in vitro tumor cell lines. FD-891 showed 2 approximately 7 times stronger activity than doxorubicin whereas FD-892 was 20 approximately 100 fold weaker than FD-891 and doxorubicin.

A new isopatulin derivative pintulin produced by Penicillium vulpinum F-4148 taxonomy, isolation, physico-chemical properties, structure and biological properties.

During our screening program of natural products from fungal metabolites for drugs effective against tumor cell lines, we discovered a new isopatulin derivative, pintulin, from the fermentation broth of Penicillium vulpinum F-4148. Pintulin shows weak activity against tumor cell lines, compared to that of adriamycin.

Lonomycins B and C, two new components of polyether antibiotics. Fermentation, isolation and characterization.

Lonomycin B (II), C44H75O14Na, m.p. 181-182 degrees C, and lonomycin C (III), C43H73O14Na, m.p 186-187 degrees C, were isolated as their sodium salts from the fermentation broth of Streptomyces ribosidificus TM-481. Their physicochemical properties demonstrated that II and III are closely related congeners of lonomycin A (I). The identical mass spectra of methyl esters of I and II indicated that II is a stereoisomer of I. On the other hand, the mass spectrum of a methyl ester of III showed a peak at m/e 810 due to M+-H2O which is smaller by 14 mass units than the maximum peak at m/e 824 due to M+-H2O of the methyl esters of I and II. This result together with the elemental analysis strongly suggested that III is a demethyl derivative of I or II. II and III are slightly less active than I in their antimicrobial activities.

Studies on the ionophorous antibiotics. XXV. The assignments of the 13C-NMR spectra of dianemycin and lenoremycin.

All the resonances observed in the 13C-NMR spectra of polyether antibiotics, dianemycin and lenoremycin (Ro 21-6150) have been assigned by the aid of selective proton decoupling experiments, T1 value measurements and biosynthetic methods as well as comparison to model compounds such as monensin, nigericin, etheromycin and carriomycin.

Stachybotrin C and parvisporin, novel neuritogenic compounds. I. Taxonomy, isolation, physico-chemical and biological properties.

Stachybotrin C and parvisporin, novel neuritogenic compounds, were isolated from the culture broth of Stachybotrys parvispora F4708. Stachybotrin C induced significant neurite outgrowth in PC12 cells and showed cell survival activity in the primary culture of cerebral cortical neurons. Parvisporin demonstrated only weak neuritogenic activity.

Structure of NG-061, a novel potentiator of nerve growth factor (NGF) isolated from Penicillium minioluteum F-4627.

The structure of NG-061, a new potentiator of nerve growth factor (NGF) isolated from Penicillium minioluteum F-4627, was determined by spectroscopic analysis and X-ray diffraction method to be phenylacetic acid 2-(2-methoxy-4-oxocyclohexa-2,4-dienylidene)-hydrazide.

A novel fungal metabolite NG-061 enhances and mimics neurotrophic effect of nerve growth factor (NGF) on neurite outgrowth in PC12 cells.

During the course of our screening program for low molecular natural products with their ability to potentiate and/or mimic neurotrophic effect of NGF, a novel fungal metabolite, phenylacetic acid hydrazide derivative NG-061 was isolated from the fermentation broth of Penicillium minioluteum F-4627. NG-061 enhanced and mimicked neurotrophic effect of NGF on neurite outgrowth in a rat pheochromocytoma cell line PC12.

Isolation and characterization of a new 12-membered macrolide FD-895.

During the course of our screening program for natural product drugs effective against multidrug resistant cells by using adriamycin resistant HL-60 cells, we have discovered a new 12 membered macrolide FD-895 in the fermentation broth of Streptomyces hygroscopicus A-9561 isolated from a soil sample collected at Iriomote Island, Okinawa prefecture, Japan. FD-895 showed stronger cytocidal activities against in vitro tumor cell lines than adriamycin. FD-895 had the same IC50 values against parent and adriamycin resistant HL-60 cells.

A novel bioactive delta lactone FD-211. Taxonomy, isolation and characterization.

During our screening program for natural product drugs effective against multidrug-resistant mammalian cells, we have discovered a new delta lactone FD-211 from the fermantation broth of Myceliophthora lutea TF-0409. FD-211 had a broad spectrum activity against cultured tumor cell lines, including adriamycin-resistant HL-60 cells.