Biology subjects

Ravenburg, C. M.

Publications and source records attributed to Ravenburg, C. M..

3 recordsLinked to original sources

bioRxiv2026

The chloroplast CLP chaperone-protease system controls the steady-state abundance of the singlet oxygen sensors EXCUTER 1 and 2

O_LIThe chloroplast CLP chaperone-protease is essential for chloroplast biogenesis. CLP substrate selection is aided by the N-recognin CLPS1 and CLPF adaptors. They interact with each other and the CLPC1 chaperone, but their specific functions are poorly understood. C_LIO_LIWe employed in vivo CLPC1 substrate-trapping in Arabidopsis by expressing a 35S:CLPC1-TRAP-STREPII transgene in wild-type (WT), clpf, clps1, and clpfclps1 to test the consequences of the loss of these adaptors on CLPC1-trapped proteins. Immunoblotting and protein half-life experiments were carried out for identified CLP substrates. C_LIO_LIExpression of the 35S:CLPC1-TRAP-STREPII in clps1cpf was embryo lethal. CLPF was trapped at a reduced level in clps1, supporting CLPS-CLPF interactions. Chloroplast 1O2 sensor EXECUTER1 (EX1) was trapped in WT and clps1 but not significantly in clpf. Steady-state protein accumulation of EX1 and its homolog EX2 increased 30-fold in the CLPC1-TRAP lines and clpr2-1, but not in clpf or clps1. In planta experiments showed that the half-life of EX1 is [~]3-fold longer in clpc1-1 than in WT, but EX1 half-life was unaffected in clpf. C_LIO_LIWe conclude that the CLP system plays a key role in EX1,2 homeostasis by keeping their intra-chloroplast concentrations low through continuous degradation, upstream of their 1O2 signaling function. C_LI

plant biology↗

bioRxiv2021

The β-amylase7 gene in Zea mays encodes a protein with structural and catalytic properties similar to Arabidopsis BAM2

Starch accumulates in the plastids of green plant tissue during the day to provide carbon for metabolism at night. Starch hydrolysis is catalyzed by members of the {beta}-amylase (BAM) family, which in Arabidopsis thaliana (At), includes nine structurally and functionally diverse members. One of these enzymes, AtBAM2, is a plastid-localized enzyme that is unique among characterized {beta}-amylases since it is tetrameric and exhibits sigmoidal kinetics. Sequence alignments show that the BAM domains of AtBAM7, a catalytically inactive, nuclear-localized transcription factor with an N-terminal DNA binding domain, and AtBAM2 are more closely related to each other than they are to any other AtBAM. Since BAM2 is found in more ancient lineages, it was hypothesized that BAM7 evolved from BAM2. However, analysis of the genomes of 48 flowering plants revealed 12 species that appear to have a BAM7 gene but lack a BAM2 gene. Upon closer inspection, these BAM7 proteins have a greater percent identity to AtBAM2 than to AtBAM7, and they share all of the AtBAM2 functional residues that BAM7 proteins normally lack. We hypothesize that these genes may encode a BAM2-like protein although they are currently annotated as BAM7-like genes. To test this hypothesis, we designed a cDNA of the short form of corn BAM7 (ZmBAM7-S) for expression in E. coli. Small Angle X-Ray Scattering data indicate that ZmBAM7-S has a tetrameric solution structure more similar to that of AtBAM2 than AtBAM1. In addition, partially purified ZmBAM7-S is catalytically active and exhibits sigmoidal kinetics. Together these data suggest that some BAM7 genes may encode a functional BAM2. Exploring and understanding {beta}-amylase gene structure could have impacts on the current annotation of genes.

biochemistry↗

bioRxiv2019

Solution structure and assembly of β-amylase 2 from Arabidopsis thaliana

Starch is a key energy storage molecule in plants that requires controlled synthesis and breakdown for effective plant growth. {beta}-amylases (BAMs) hydrolyze starch into maltose to help meet the metabolic needs of the plant. In the model plant, Arabidopsis thaliana, there are nine BAMs which have apparently distinct functional and domain structures, although the functions of only a few of the BAMs are known and there are no 3-D structures of BAMs from this organism. Recently, AtBAM2 was proposed to form a tetramer based on chromatography and activity assays of mutants, however there was no direct observation of this tetramer. We collected small-angle X-ray scattering data on AtBAM2 and N-terminal mutants to describe the structure and assembly of the tetramer. Comparison of the scattering of the AtBAM2 tetramer to data collected using the sweet potato (Ipomoea batatas) BAM5, which is also reported to form a tetramer, showed there were differences in the overall assembly. Analysis of N-terminal truncations of AtBAM2 identified a loop sequence found only in BAM2 orthologs that appears to be critical for AtBAM2 tetramer assembly as well as activity.

biochemistry↗