Biology subjects

Munro, K.

Publications and source records attributed to Munro, K..

3 recordsLinked to original sources

bioRxiv2023

Structure and function of a hexameric cyanophycin synthetase 2

Cyanophycin is a natural polymer composed of a poly-aspartate backbone with arginine attached to each of the aspartate sidechains. Produced by a wide range of bacteria, which mainly use it as a store of fixed nitrogen, it has many promising industrial applications. Cyanophycin can be synthesized from the amino acids Asp and Arg by the widespread cyanophycin synthetase 1 (CphA1), or from the dipeptide {beta}-Asp-Arg by the cyanobacterial enzyme cyanophycin synthetase 2 (CphA2). CphA2 enzymes display a range of oligomeric states, from dimers to dodecamers. Recently, the crystal structure of a CphA2 dimer was solved but could not be obtained in complex with substrate. Here, we report cryo-EM structures of the hexameric CphA2 from Stanieria sp. at ~2.8 [A] resolution, both with and without ATP and cyanophycin. The structures show a trimer-of-dimers hexameric architecture, and substrate-binding interactions that are similar to those of CphA1. Mutagenesis experiments demonstrate the importance of several conserved substrate-binding residues. We also find that a Q416A/R528G double mutation prevents hexamer formation and use this double mutant to show that hexamerization augments the rate of cyanophycin synthesis. Together, these results increase our mechanistic understanding of how an interesting green polymer is biosynthesized.

biochemistry↗

bioRxiv2023

Arabidopsis calmodulin-like proteins CML13 and CML14 interact with proteins that have IQ domains

In response to Ca2+ signals, the evolutionarily-conserved Ca2+ sensor calmodulin (CaM) regulates protein targets via direct interaction. Plants possess many CaM-like (CML) proteins, but their binding partners and functions are mostly unknown. Here, using Arabidopsis CML13 as bait in a yeast two-hybrid screen, we isolated putative targets from three, unrelated protein families, namely, IQD proteins, calmodulin-binding transcriptional activators (CAMTAs), and myosins, all of which possess tandem isoleucine-glutamine (IQ) structural domains. Using the split-luciferase complementation assay in planta and the yeast 2-hybrid system, CML13 and CML14 showed a preference for interaction with tandem over single IQ domains. Relative to CaM, CML13 and CML14 displayed weaker signals when tested with the non-IQ, CaM-binding domain of glutamate decarboxylase or the single IQ domains of CNGC20 (cyclic-nucleotide gated channel-20) or IQM1 (IQ motif protein1). We examined IQD14 as a representative tandem IQ-protein and found that only CaM, CML13, and CML14 interacted with IQD14 among 12 CaM/CMLs tested. CaM, CML13, and CML14 bound in vitro to IQD14 in the presence or absence of Ca2+. Binding affinities were in the nM range and were higher when two tandem IQ domains from IQD14 were present. Green fluorescent protein-tagged versions of CaM, CML13, and CML14 localized to both the cytosol and nucleus in plant cells but were partially relocalized to the microtubules when co-expressed with IQD14 tagged with mCherry. These and other data are discussed in the context of possible roles for these CMLs in gene regulation via CAMTAs and cytoskeletal activity via myosins and IQD proteins.

plant biology↗

bioRxiv2020

Gut microbiome composition predicts summer core range size in a generalist and specialist ungulate

The gut microbiome of animals varies by age, diet, and habitat, and directly influences individual health. Similarly, variation in an individuals home range can lead to differences in feeding strategies and fitness. Ungulates (hooved mammals) exhibit species-specific microbiomes and habitat use patterns: here, we combined gut microbiome and movement data to assess relationships between space use and the gut microbiome in a specialist and a generalist ungulate. We captured and GPS radiocollared 24 mountain goats (Oreamnos americanus) and 34 white-tailed deer (Odocoileus virginianus). We collected fecal samples and conducted high-throughput sequencing of the 16S rRNA gene. Using GPS data, we estimated core (50%) and home range (95%) sizes and calculated proportional use for several important habitat types. We generated metrics related to gut diversity and key bacterial ratios. We hypothesize that larger Firmicutes to Bacteroides ratios confer body size or fat advantages that allow for larger home ranges, and that relationships between gut diversity and disproportionate habitat use is stronger in mountain goats due to their restricted niche relative to white-tailed deer. Firmicutes to Bacteroides ratios were positively correlated with core range area in both species. Mountain goats exhibited a negative relationship between gut diversity and use of two key habitat types (treed areas and escape terrain), whereas no relationships were detected in white-tailed deer. This is the first study to relate core range size to the gut microbiome in wild ungulates and is an important proof of concept that advances the information that can be gleaned from non-invasive sampling.

molecular biology↗