Characterisation of Modular Polyketide Synthases Designed to Make Pentaene Analogues of Amphotericin B.

Glycosylated polyene macrolides are important antifungal agents that are produced by many actinomycete species. Development of new polyenes may deliver improved antibiotics. Here, was genetically re-programmed to synthesise pentaene analogues of the heptaene amphotericin B.

Biosynthesis of a new skyllamycin in : a cytochrome P450 forms an epoxide in the cinnamoyl chain.

Activation of a silent gene cluster in leads to synthesis of a cinnamoyl-containing non-ribosomal peptide (CCNP) that is related to skyllamycins. This novel CCNP was isolated and its structure was interrogated using mass spectrometry and nuclear magnetic resonance spectroscopy. The isolated compound is an oxidised skyllamycin A in which an additional oxygen atom is incorporated in the cinnamoyl side-chain in the form of an epoxide.

Thermococcus kodakarensis TK0353 is a novel AP lyase with a new fold.

Hyperthermophilic organisms thrive in extreme environments prone to high levels of DNA damage. Growth at high temperature stimulates DNA base hydrolysis resulting in apurinic/apyrimidinic (AP) sites that destabilize the genome. Organisms across all domains have evolved enzymes to recognize and repair AP sites to maintain genome stability.

A Biomimetic Polymer for the Extraction and Purification of Superior Analogues of Amphotericin B.

Amphotericin B has been an essential drug in the fight against leishmaniasis and fungal pathogens for decades, and has more recently gained attention for the very limited microbial resistance displayed against it. However, its toxicity has restricted its use to only the most severe cases of disease, and attempts to reduce these ill effects via formulation have had only minor success. Genetic engineering has allowed the development of superior amphotericin analogues, notably 16-descarboxyl-16-methyl amphotericin B (MeAmB), which shows a ten-fold reduction in toxicity in addition to a slight improvement in therapeutic activity.

New Glycosylated Polyene Macrolides: Refining the Ore from Genome Mining.

Glycosylated polyene macrolides include effective antifungal agents, such as pimaricin, nystatin, candicidin, and amphotericin B. For the treatment of systemic mycoses, amphotericin B has been described as a gold-standard antibiotic because of its potent activity against a broad spectrum of fungal pathogens, which do not readily become resistant. However, amphotericin B has severe toxic side effects, and the development of safer alternatives remains an important objective.

Structural analysis of P450 AmphL from Streptomyces nodosus provides insights into substrate selectivity of polyene macrolide antibiotic biosynthetic P450s.

AmphL is a cytochrome P450 enzyme that catalyzes the C8 oxidation of 8-deoxyamphotericin B to the polyene macrolide antibiotic, amphotericin B. To understand this substrate selectivity, we solved the crystal structure of AmphL to a resolution of 2.0 Å in complex with amphotericin B and performed molecular dynamics (MD) simulations.

Capillary Electrophoresis-Based Functional Genomics Screening to Discover Novel Archaeal DNA Modifying Enzymes.

It has been predicted that 30 to 80% of archaeal genomes remain annotated as hypothetical proteins with no assigned gene function. Further, many archaeal organisms are difficult to grow or are unculturable. To overcome these technical and experimental hurdles, we developed a high-throughput functional genomics screen that utilizes capillary electrophoresis (CE) to identify nucleic acid modifying enzymes based on activity rather than sequence homology.

Supporting Newly Arrived Migrant Mothers: A Pilot Health Literacy Intervention.

Experiencing migration can create or exacerbate vulnerability to ill health, particularly during pregnancy and new motherhood. Providing a culturally appropriate health literacy intervention to new migrant families may increase social support and the skills and confidence to access health care services and information. This study developed and piloted a health literacy intervention, in the form of culturally redesigned new parent classes, in a culturally diverse location in Australia.

Nucleosome Core Particles Lacking H2B or H3 Tails Are Altered Structurally and Have Differential Base Excision Repair Fingerprints.

A recently discovered post-translational modification of histone proteins is the irreversible proteolytic clipping of the histone N-terminal tail domains. This modification is involved in the regulation of various biological processes, including the DNA damage response. In this work, we used chemical footprinting to characterize the structural alterations to nucleosome core particles (NCPs) that result from a lack of a histone H2B or H3 tail.

Comparison of the Base Excision and Direct Reversal Repair Pathways for Correcting 1,-Ethenoadenine in Strongly Positioned Nucleosome Core Particles.

1,-ethenoadenine (εA) is a mutagenic lesion and biomarker observed in numerous cancerous tissues. Two pathways are responsible for its repair: base excision repair (BER) and direct reversal repair (DRR). Alkyladenine DNA glycosylase (AAG) is the primary enzyme that excises εA in BER, generating stable intermediates that are processed by downstream enzymes.

Access to appropriate health care for non-English speaking migrant families with a newborn/young child: a systematic scoping literature review.

Background: Recently arrived culturally and linguistically diverse migrant mothers in Western Industrialised Nations are less likely to access health care and are more likely to report negative healthcare experiences than more established migrant or non-migrant populations. This is particularly an issue in Australia where nearly half of all Australians were born overseas or have at least one parent born overseas.

Methods: A systematic scoping review was conducted to identify a) the main enablers and barriers to accessing appropriate health care for migrant families with a new baby/young child who speak a language other than English, and b) the effectiveness of interventions that have been tested to improve access to appropriate health care for this group.

Chromatin and other obstacles to base excision repair: potential roles in carcinogenesis.

DNA is comprised of chemically reactive nucleobases that exist under a constant barrage from damaging agents. Failure to repair chemical modifications to these nucleobases can result in mutations that can cause various diseases, including cancer. Fortunately, the base excision repair (BER) pathway can repair modified nucleobases and prevent these deleterious mutations.

Initiating base excision repair in chromatin.

The base excision repair (BER) pathway removes modified nucleobases that can be deleterious to an organism. BER is initiated by a glycosylase, which finds and removes these modified nucleobases. Most of the characterization of glycosylase activity has been conducted in the context of DNA oligomer substrates.

Phase I trial of selenium plus chemotherapy in gynecologic cancers.

Purpose: Preclinical studies performed in our laboratory have shown that high-dose selenium inhibits the development of carboplatin drug resistance in an ovarian cancer mouse xenograft model. Based on these data, as well as the potential serious toxicities of supranutritional doses of selenium, a phase I trial of a combination of selenium/carboplatin/paclitaxel was designed to determine the maximum tolerated dose, safety, and effects of selenium on carboplatin pharmacokinetics in the treatment of chemo-naive women with gynecologic cancers. Correlative studies were performed to identify gene targets of selenium.

New insights into polyene macrolide biosynthesis in Couchioplanes caeruleus.

Couchioplanes caeruleus DSM43634 synthesises 67-121C, an aromatic heptaene macrolide that contains a mannosyl-mycosaminyl disaccharide. An improved draft genome sequence was used to obtain the biosynthetic gene cluster for this antifungal. Bioinformatic analysis of the polyketide synthase indicated that extension modules 7 and 8 contain A-type ketoreductase and dehydratase domains.

Engineered biosynthesis and characterisation of disaccharide-modified 8-deoxyamphoteronolides.

Several polyene macrolides are potent antifungal agents that have severe side effects. Increased glycosylation of these compounds can improve water solubility and reduce toxicity. Three extending glycosyltransferases are known to add hexoses to the mycosaminyl sugar residues of polyenes.

A Model of the Development of Cisplatin Resistance in Human Small Cell Lung Cancer Xenografts.

Background/aim: To study the prevention of chemotherapy resistance, we have previously designed models of drug-resistant ovarian cancer. We here report an in vivo model of cisplatin-resistant small cell lung cancer (SCLC).

Materials And Methods: Mice bearing H526 SCLC xenografts received intraperitoneal pretreatment with a sub-effective cisplatin dose (0.

Corrigendum to "Dissecting complex polyketide biosynthesis" [Comput Struct Biotechnol J 3(4) (2012) 1-8].

[This corrects the article DOI: 10.5936/csbj.201210010.

Polyene macrolide biosynthesis in streptomycetes and related bacteria: recent advances from genome sequencing and experimental studies.

The polyene macrolide group includes important antifungal drugs, to which resistance does not arise readily. Chemical and biological methods have been used in attempts to make polyene antibiotics with fewer toxic side effects. Genome sequencing of producer organisms is contributing to this endeavour, by providing access to new compounds and by enabling yield improvement for polyene analogues obtained by engineered biosynthesis.

Exploiting the genome sequence of Streptomyces nodosus for enhanced antibiotic production.

The genome of the amphotericin producer Streptomyces nodosus was sequenced. A single scaffold of 7,714,110 bp was obtained. Biosynthetic genes were identified for several natural products including polyketides, peptides, siderophores and terpenes.

Role of polyol moiety of amphotericin B in ion channel formation and sterol selectivity in bilayer membrane.

Amphotericin B (AmB) is a polyene macrolide antibiotic widely used to treat mycotic infections. In this paper, we focus on the role of the polyol moiety of AmB in sterol selectivity using 7-oxo-AmB, 7α-OH-AmB, and 7β-OH-AmB. The 7-OH analogs were prepared from 7-oxo-AmB.

Prevention of carboplatin-induced resistance in human ovarian tumor xenografts by selenite.

Background/aim: We have been exploring a prevention approach to the problem of drug resistance which develops during ovarian cancer chemotherapy. We have previously described an in vivo model of the development of resistance to the chemotherapy drug cisplatin in xenografts, and the prevention of this resistance by selenium compounds. However, a different platinum-based drug, carboplatin, is frequently utilized in ovarian cancer treatment.

Engineered biosynthesis of disaccharide-modified polyene macrolides.

Recent work has uncovered genes for two glycosyltransferases that are thought to catalyze mannosylation of mycosaminyl sugars of polyene macrolides. These two genes are nypY from Pseudonocardia sp. strain P1 and pegA from Actinoplanes caeruleus.

Versatility of enzymes catalyzing late steps in polyene 67-121C biosynthesis.

Actinoplanes caeruleus produces 67-121C, a heptaene macrolide modified with a D-mannosyl-D-mycosaminyl disaccharide. Draft genome sequencing revealed genes encoding mycosaminyltransferase, mycosamine synthase, a cytochrome P450 that modifies the macrolactone core, and the extending mannosyltransferase. Only the mycosamine synthase and P450 were active in the biosynthesis of amphotericins in Streptomyces nodosus, the amphotericin producer.