A prevalent peptide-binding domain guides ribosomal natural product biosynthesis - PubMed (original) (raw)

A prevalent peptide-binding domain guides ribosomal natural product biosynthesis

Brandon J Burkhart et al. Nat Chem Biol. 2015 Aug.

Abstract

Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a rapidly growing class of natural products. RiPP precursor peptides can undergo extensive enzymatic tailoring to yield structurally and functionally diverse products, and their biosynthetic logic makes them attractive bioengineering targets. Recent work suggests that unrelated RiPP-modifying enzymes contain structurally similar precursor peptide-binding domains. Using profile hidden Markov model comparisons, we discovered related and previously unrecognized peptide-binding domains in proteins spanning the majority of known prokaryotic RiPP classes, and we named this conserved domain the RiPP precursor peptide recognition element (RRE). Through binding studies we verified RRE's roles for three distinct RiPP classes: linear azole-containing peptides, thiopeptides and lasso peptides. Because numerous RiPP biosynthetic enzymes act on peptide substrates, our findings have powerful predictive value as to which protein(s) drive substrate binding, thereby laying a foundation for further characterization of RiPP biosynthetic pathways and the rational engineering of new peptide-binding activities.

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Conflict of interest statement

Competing financial interests

The authors declare no competing financial interests.

Figures

Figure 1

Figure 1. Overview of RiPP biosynthesis and the TOMM subclass

(a) A generic RiPP biosynthetic gene cluster is displayed. The precursor peptide is composed of a leader peptide (LP) and core. While the LP contains binding motifs for the modifying enzymes, the core contains residues that undergo enzymatic processing to diverse functional groups. After removal of the LP and any additional tailoring processes, the mature RiPP is exported. (b) One RiPP biosynthetic class, the thiazole/oxazole-modified microcins (TOMMs), installs azoline/azole heterocycles. TOMMs arise from the action of an ATP-dependent cyclodehydratase (C and D proteins) and a flavin mononucleotide (FMN)-dependent dehydrogenase (B protein). X = S or O; R = H or CH3.

Figure 2

Figure 2. Structural comparison of four RiPP modifying enzymes

Shown are structurally equivalent sections of RiPP biosynthetic proteins involved in the biosynthesis of (a) the Trojan horse antibiotic microcin C7 (MccB, an adenylating enzyme [PDB entry 3H9J]), (b) antitumor cyanobactins (LynD, a cyclodehydratase [4V1T]), (c) the lantibiotic nisin (NisB, dehydratase [4WD9]), and (d) the bacterial dehydrogenase cofactor PQQ (PqqD, rSAM-associated [3G2B]). The purple β-sheets and cyan α-helices constitute a conserved

R

iPP precursor peptide

r

ecognition

e

lement (RRE). In (a–c), the precursor peptide is shown in yellow stick format. Dashed lines indicate missing electron density.

Figure 3

Figure 3. RREs are present in diverse RiPP biosynthetic proteins

RREs were found in a myriad of RiPP biosynthetic proteins using HHpred to search for PqqD homology (thick black bars). Solid lines represent protein sequences with N/_C_-termini labeled as “N” and “C.” Colored sections indicate the annotations for the conserved domains identified by the Conserved Domain Database. Asterisks denote RRE assignments based solely on HHpred findings. Abbreviations: LAP, linear azole-containing peptide; PQQ, pyrroloquinoline quinone; rSAM, radical SAM. More details can be found in Supplementary Table 3.

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