Biology subjects

Weir, E.

Publications and source records attributed to Weir, E..

4 recordsLinked to original sources

bioRxiv2026

Structure and activity of a class II lanthipeptide from a thermophilic bacterium

Lanthipeptides represent a large group of ribosomally synthesized and post-translationally modified peptides (RiPPs). They offer promising avenues for discovering new antibacterial and antifungal agents. Here, we identify and structurally analyze the product of the tla BGC, which encodes a class II lanthipeptide in the thermophilic bacterium Thermoactinomyces sp. DSM 45891. Co-expression of the lanthipeptide synthetase TlaM with its substrates in Escherichia coli resulted in modification of the two precursor peptides TlaA1 and TlaA2, which share 58% sequence identity. TlaA1 was dehydrated up to seven times, with the major products having undergone five and six dehydrations, whereas TlaA2 was dehydrated seven times. In both peptides, four thioether rings were formed with two overlapping DL-(methyl)lanthionine rings at the C-terminus. Both peptides also contain two non-overlapping DL-methyllanthionines near the N-terminus and in the center of the peptide. These peptides deviate from the general rule of stereoselective LL-(methyl)lanthionine formation from a Dhx-Dhx-Xxx-Xxx-Cys motif (Dhx = dehydroalanine or dehydrobutyrine). AspN-cleaved TlaM-modified TlaA1 displayed anti-microbial activity against a subset of bacteria including Gram-negative ESKAPE pathogens. We named the lantibiotic thermolanthin. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=175 SRC="FIGDIR/small/716199v2_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@11b6dd0org.highwire.dtl.DTLVardef@1c6a68borg.highwire.dtl.DTLVardef@e026f4org.highwire.dtl.DTLVardef@1d7d6b_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2026

Structure, biosynthesis, and bioactivity of nostolysamides

A recent genome mining study identified class II lanthipeptides encoded in Nostoc punctiforme PCC 73102 that contain acyl groups conjugated to Lys side chains. The structure and bioactivity of these peptides, named nostolysamides, were not determined. In this study, we produced the nostolysamides by co-expression of the NpuA precursor peptide with an N-terminal SUMO tag with the class II lanthipeptide synthetase NpuM in Escherichia coli. All four lanthionine and methyllanthionine residues were shown to have the DL configuration by Marfeys analysis. Tandem mass spectrometry and mutagenesis studies indicate an N-terminal non-overlapping methyllanthionine ring and three overlapping rings at the C-terminus for which the most likely ring pattern is proposed. The NpuM lanthipeptide synthetase is a member of the ProcM-clade and catalyzes ring formation with both C-to-N and N-to-C directionality. After removal of the leader peptide, the resulting lanthipeptide exhibits antibacterial as well as antifungal activity against Candida species by disrupting cell membranes. Antibacterial activity is shown not to involve lipid II. The biosynthetic gene cluster also encodes an acetyltransferase NpuN that transfers long chain acyl groups to the side chain of a Lys residue in position 1 of the precursor peptide. In vitro studies of NpuN show relatively broad substrate specificity with NpuN conjugating various acyl groups from acyl-CoA substrates to Lys1 in the nostolysamides. The acylation did not appreciably change the antifungal and antimicrobial activity of nostolysamide showing that it is not required for these activities.

biochemistry↗

bioRxiv2019

ER transmembrane protein TMTC3 contributes to O-mannosylation of E-cadherin, Cellular Adherence and Embryonic Gastrulation

Protein glycosylation plays essential roles in protein structure, stability and activity such as cell adhesion. The cadherin superfamily of adhesion molecules carry O-linked mannose glycans at conserved sites and it was recently demonstrated that the TMTC1-4 genes contribute to the addition of these O-linked mannoses. Here, biochemical, cell biological and organismal analysis was used to determine that TMTC3 supports the O-mannosylation of E-cadherin, cellular adhesion and embryonic gastrulation. Using genetically engineered cells lacking all four TMTC genes, overexpression of TMTC3 rescued O-linked glycosylation of E-cadherin and cell adherence. The knockdown of the Tmtcs in Xenopus laevis embryos caused a delay in gastrulation that was rescued by the addition of human TMTC3. Mutations in TMTC3 have been linked to neuronal cell migration diseases including Cobblestone lissencephaly. Analysis of TMTC3 mutations associated with Cobblestone lissencephaly found that three of the variants exhibit reduced stability and missence mutations were unable to complement TMTC3 rescue of gastrulation in Xenopus embryo development. Our study demonstrates that TMTC3 regulates O-linked glycosylation and cadherin-mediated adherence, providing insight into its effect on cellular adherence and migration, as well the basis of TMTC3-associated Cobblestone lissencephaly.

cell biology↗

bioRxiv2019

A meiosis-specific AAA+ assembly reveals repurposing of ORC during budding yeast gametogenesis

ORC (Orc1-6) is an AAA+ complex that loads the AAA+ MCM helicase to replication origins. Orc1, a subunit of ORC, functionally interacts with budding yeast Pch2, a meiosis-specific AAA+ protein. Pch2 regulates several chromosomal events of gametogenesis, but mechanisms that dictate Pch2 function remain poorly understood. We demonstrate that ORC directly interacts with an AAA+ Pch2 hexamer. The ORC-Pch2 assembly is established without Cdc6, a factor crucial for ORC-MCM binding. Biochemical analysis suggests that Pch2 utilizes ORCs Cdc6-binding interface and employs its non-enzymatic NH2-terminal domain and AAA+ core to engage ORC. In contrast to phenotypes observed upon Orc1 impairment, nuclear depletion of other subunits of ORC does not lead to Pch2-like phenotypes, indicating that ORC integrity per se is not required to support Pch2 function. We thus reveal functional interplay between Pch2 and ORC, and uncover the repurposing of ORC to establish a non-canonical and meiosis-specific AAA+ assembly.

biochemistry↗