Biology subjects

Krieger, I. V.

Publications and source records attributed to Krieger, I. V..

3 recordsLinked to original sources

bioRxiv2023

Bacillus subtilis YisK possesses oxaloacetate decarboxylase activity and exhibits Mbl-dependent localization

YisK is an uncharacterized protein in Bacillus subtilis previously shown to interact genetically with the elongasome protein Mbl. YisK overexpression leads to cell widening and lysis, phenotypes that are dependent on mbl and suppressed by mbl mutations. In the present work we characterize YisKs localization, structure, and enzymatic activity. We show that YisK localizes in a punctate and/or punctate-helical pattern that depends on Mbl, and that YisK interacts directly with another elongasome protein, FtsE. YisK belongs to the fumarylacetoacetate hydrolase (FAH) superfamily and crystal structures revealed close structural similarity to two oxaloacetate (OAA) decarboxylases: human mitochondrial FAHD1 and Corynebacterium glutamicum Cg1458. We demonstrate that YisK can also catalyze the decarboxylation of OAA (Km = 134 {micro}M, Kcat = 31 min-1). A catalytic dead variant (YisK E148A, E150A) retains wild-type localization and still widens cells following overexpression, indicating these activities are not dependent on YisK catalysis. Conversely, a non-localizing variant (YisK E30A) retains wild-type enzymatic activity in vitro, but no longer widens cells following overexpression. Together these results suggest YisK may be subject to spatial regulation that depends on the cell envelope synthesis machinery. IMPORTANCEThe elongasome is a protein complex that guides lengthwise growth in some bacteria. We previously showed that in B. subtilis, overexpression of an uncharacterized enzyme (YisK), perturbed function of the actin-like elongasome protein Mbl. Here we show that YisK exhibits Mbl-dependent localization and interacts directly with another component of the elongasome, FtsE. Through biochemical and structural characterization, we demonstrate that like its mitochondrial homolog FAHD1, YisK can catalyze the decarboxylation of the oxaloacetate to pyruvate and CO2. YisK is the first example of an enzyme implicated in central carbon metabolism with subcellular localization that depends on Mbl.

microbiology↗

bioRxiv2021

Structural anatomy of C1 domain interactions with DAG and other agonists

Diacylglycerol (DAG) is a versatile lipid whose 1,2-sn-stereoisomer serves both as second messenger in signal transduction pathways that control vital cellular processes, and as metabolic precursor for downstream signaling lipids such as phosphatidic acid1,2. DAG effector proteins compete for available lipid using conserved homology 1 (C1) domains as DAG-sensing modules. Yet, how C1 domains recognize and capture DAG in the complex environment of a biological membrane has remained unresolved for the 40 years since the discovery of Protein Kinase C (PKC)3 as the first member of the DAG effector cohort. Herein, we report the first high-resolution crystal structures of a C1 domain (C1B from PKC{delta}) complexed to DAG and to each of four potent PKC agonists that produce different biological readouts and that command intense therapeutic interest. This structural information details the mechanisms of stereospecific recognition of DAG by the C1 domains, the functional properties of the lipid-binding site, and the identities of the key residues required for the recognition and capture of DAG and exogenous agonists. Moreover, the structures of the five C1 domain complexes provide the high-resolution guides for the design of agents that modulate the activities of DAG effector proteins.

biophysics↗

bioRxiv2021

A class I hydrophobin in Trichoderma virens influences plant-microbe interactions through enhancement of enzyme activity and MAMP recognition

The filamentous fungus, Trichoderma virens, is a well-known mycoparasitic plant symbiont, valued for its biocontrol capabilities. T. virens initiates a symbiotic relationship with a plant host through the colonization of its roots. To achieve colonization, the fungus must communicate with the host and evade its innate defenses. Hydrophobins from Trichoderma spp. have previously been demonstrated to be involved in colonization of host roots. In this study, the class I hydrophobin, HFB9A from T. virens was characterized for a potential role in root colonization. {Delta}hfb9a gene deletion mutants colonized less than the wild-type strain, were unable to induce systemic resistance against Colletotrichum graminicola, and showed a reduction in the activity of its cell wall degrading enzymes. The purified HFB9A protein was able to complement the enzyme activity of mutant culture filtrates as well as enhance the activity of commercially sourced cellulase. When exogenously applied to Arabidopsis plants, HFB9A protein induced phosphorylation of AtMAPK3/6, suggesting that it functions as a microbe-associated molecular pattern.

plant biology↗