Motivation: In 2001 and 2002, we published two papers (Bioinformatics, 17, 282--283, Bioinformatics, 18, 77--82) describing an ultrafast protein sequence clustering program called cd-hit. This program can efficiently cluster a huge protein database with millions of sequences. However, the applications of the underlying algorithm are not limited to only protein sequences clustering, here we present several new programs using the same algorithm including cd-hit-2d, cd-hit-est and cd-hit-est-2d. Cd-hit-2d compares two protein datasets and reports similar matches between them; cd-hit-est clusters a DNA/RNA sequence database and cd-hit-est-2d compares two nucleotide datasets. All these programs can handle huge datasets with millions of sequences and can be hundreds of times faster than methods based on the popular sequence comparison and database search tools, such as BLAST.

Availability: http://cd-hit.org

Contact: [email protected]

Cd-hit: a fast program for clustering and comparing large sets of protein or nucleotide sequences

The alignment of sequencing reads against a protein reference database is a major computational bottleneck in metagenomics and data-intensive evolutionary projects. Although recent tools offer improved performance over the gold standard BLASTX, they exhibit only a modest speedup or low sensitivity. We introduce DIAMOND, an open-source algorithm based on double indexing that is 20,000 times faster than BLASTX on short reads and has a similar degree of sensitivity.

Fast and sensitive protein alignment using DIAMOND

This review is an updated and expanded version of two prior reviews that were published in this journal in 1997 and 2003. In the case of all approved agents the time frame has been extended to include the 251/2 years from 01/1981 to 06/2006 for all diseases worldwide and from 1950 (earliest so far identified) to 06/2006 for all approved antitumor drugs worldwide. We have continued to utilize our secondary subdivision of a “natural product mimic” or “NM” to join the original primary divisions. From the data presented, the utility of natural products as sources of novel structures, but not necessarily the final drug entity, is still alive and well. Thus, in the area of cancer, over the time frame from around the 1940s to date, of the 155 small molecules, 73% are other than “S” (synthetic), with 47% actually being either natural products or directly derived therefrom. In other areas, the influence of natural product structures is quite marked, with, as expected from prior information, the antiinfective area ...

/pdf/natural-products-as-sources-of-new-drugs-over-the-last-25-24stxjn5tu.pdf

Natural Products as Sources of New Drugs over the Last 25 Years

https://pubs.rsc.org/en/content/articlepdf/2019/NP/C8NP00092A

Marine natural products.

This contribution is a completely updated and expanded version of the four prior analogous reviews that were published in this journal in 1997, 2003, 2007, and 2012. In the case of all approved therapeutic agents, the time frame has been extended to cover the 34 years from January 1, 1981, to December 31, 2014, for all diseases worldwide, and from 1950 (earliest so far identified) to December 2014 for all approved antitumor drugs worldwide. As mentioned in the 2012 review, we have continued to utilize our secondary subdivision of a “natural product mimic”, or “NM”, to join the original primary divisions and the designation “natural product botanical”, or “NB”, to cover those botanical “defined mixtures” now recognized as drug entities by the U.S. FDA (and similar organizations). From the data presented in this review, the utilization of natural products and/or their novel structures, in order to discover and develop the final drug entity, is still alive and well. For example, in the area of cancer, over t...

Natural Products as Sources of New Drugs from 1981 to 2014

X-ray single-crystal diffraction (SCD) analysis has the intrinsic limitation that the target molecules must be obtained as single crystals. Here we report a protocol for SCD analysis that does not require the crystallization of the sample. In our method, tiny crystals of porous complexes are soaked in a solution of the target, such that the complexes can absorb the target molecules. Crystallographic analysis clearly determines the absorbed guest structures along with the host frameworks. Because the SCD analysis is carried out on only one tiny crystal of the complex, the required sample mass is of the nanogram–microgram order. We demonstrate that as little as about 80 nanograms of a sample is enough for the SCD analysis. In combination with high-performance liquid chromatography, our protocol allows the direct characterization of multiple fractions, establishing a prototypical means of liquid chromatography SCD analysis. Furthermore, we unambiguously determined the structure of a scarce marine natural product using only 5 micrograms of the compound. Chemists need reliable methods to analyse and determine molecular structures. Nuclear magnetic resonance (NMR) and mass spectrometry are indispensable tools in daily chemical research for rapidly analysing molecular structures, but, strictly speaking, they provide only speculative molecular structures that are sometimes assigned incorrectly. However, X-ray SCD provides direct structural information at the atomic level and is recognized as the most reliable structure determination method 1–3 . Unfortunately, X-ray SCD has some critical limitations. First, the crystallization of samples before measurement can not be automated and usually requires a time-consuming trial-and-error procedure. Second, the method is in principle not applicable to non-crystalline molecules. In this Article, we describe an advance in crystallographic analysis based on a new X-ray analysis protocol that does not require the crystallization of the sample molecules themselves. Our idea is to use networked porous metal complexes 4–7 as ‘crystalline sponges’ 8 . Owing to the high molecular-recognition ability of the pores, the crystalline sponges can absorb target sample molecules from their solution into the pores, rendering the incoming molecules regularly ordered in the crystal. Accordingly, the molecular structure of the absorbed guest will be displayed, along with the host framework, by the crystallographic analysis of the networked porous complexes. We emphasize that even trace amounts of samples (,0.1mg) can be analysed by this method because the experiment can be performed with only one tiny crystal (,0.1 mm to a side). In the following discussion, we thus describe the crystallographic analysis of non-crystalline compounds and nanogram–microgram-scale X-ray crystallography based on our method. The great advantage of trace-amount X-ray analysis is particularly emphasized by its application to liquid chromatography SCD analysis (see below), where high-performance liquid chromatography (HPLC) fractions are directly collected by the crystalline sponge and analysed by X-ray crystallography. Furthermore, we successfully determine the structure of a scarce marine natural product, miyakosyne A, including the absolute configuration of its chiral centre, which could not be determined by conventional chemical and spectroscopic methods. X-ray crystallography of liquid samples

X-ray analysis on the nanogram to microgram scale using porous complexes

Bacterial symbionts have long been suspected to be the true producers of many drug candidates isolated from marine invertebrates. Sponges, the most important marine source of biologically active natural products, have been frequently hypothesized to contain compounds of bacterial origin. This symbiont hypothesis, however, remained unproven because of a general inability to cultivate the suspected producers. However, we have recently identified an uncultured Pseudomonas sp. symbiont as the most likely producer of the defensive antitumor polyketide pederin in Paederus fuscipes beetles by cloning the putative biosynthesis genes. Here we report closely related genes isolated from the highly complex metagenome of the marine sponge Theonella swinhoei, which is the source of the onnamides and theopederins, a group of polyketides that structurally resemble pederin. Sequence features of the isolated genes clearly indicate that it belongs to a prokaryotic genome and should be responsible for the biosynthesis of almost the entire portion of the polyketide structure that is correlated with antitumor activity. Besides providing further proof for the role of the related beetle symbiont-derived genes, these findings raise intriguing ecological and evolutionary questions and have important general implications for the sustainable production of otherwise inaccessible marine drugs by using biotechnological strategies.

https://www.pnas.org/content/pnas/101/46/16222.full.pdf

Antitumor polyketide biosynthesis by an uncultivated bacterial symbiont of the marine sponge Theonella swinhoei

Cultivated bacteria such as actinomycetes are a highly useful source of biomedically important natural products. However, such ‘talented’ producers represent only a minute fraction of the entire, mostly uncultivated, prokaryotic diversity. The uncultured majority is generally perceived as a large, untapped resource of new drug candidates, but so far it is unknown whether taxa containing talented bacteria indeed exist. Here we report the single-cell- and metagenomics-based discovery of such producers. Two phylotypes of the candidate genus ‘Entotheonella’ with genomes of greater than 9 megabases and multiple, distinct biosynthetic gene clusters co-inhabit the chemically and microbially rich marine sponge Theonella swinhoei. Almost all bioactive polyketides and peptides known from this animal were attributed to a single phylotype. ‘Entotheonella’ spp. are widely distributed in sponges and belong to an environmental taxon proposed here as candidate phylum ‘Tectomicrobia’. The pronounced bioactivities and chemical uniqueness of ‘Entotheonella’ compounds provide significant opportunities for ecological studies and drug discovery.

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Shigeki Matsunaga

Papers

X-ray analysis on the nanogram to microgram scale using porous complexes

Antitumor polyketide biosynthesis by an uncultivated bacterial symbiont of the marine sponge Theonella swinhoei

An environmental bacterial taxon with a large and distinct metabolic repertoire

Bioactive marine metabolites. Part 16. Calyculin A. A novel antitumor metabolite from the marine sponge Discodermia calyx

Bioactive sponge peptides