Abstract
Biotin protein ligase is universal in three domains of life. The paradigm version of BPL is the Escherichia coli BirA that is also a repressor for the biotin biosynthesis pathway. Streptococcus suis, a leading bacterial agent for swine diseases, seems to be an increasingly-important opportunistic human pathogen. Unlike the scenario in E. coli, S. suis lacks the de novo biotin biosynthesis pathway. In contrast, it retains a bioY, a biotin transporter-encoding gene, indicating an alternative survival strategy for S. suis to scavenge biotin from its inhabiting niche. Here we report functional definition of S. suis birA homologue. The in vivo functions of the birA paralogue with only 23.6% identity to the counterpart of E. coli, was judged by its ability to complement the conditional lethal mutants of E. coli birA. The recombinant BirA protein of S. suis was overexpressed in E. coli, purified to homogeneity and verified with MS. Both cellulose TLC and MALDI-TOFF-MS assays demonstrated that the S. suis BirA protein catalyzed the biotinylation reaction of its acceptor biotin carboxyl carrier protein. EMSA assays confirmed binding of the bioY gene to the S. suis BirA. The data defined the first example of the bifunctional BirA ligase/repressor in Streptococcus.
Biotin (vitamin H) is one of two known sulfur-containing fatty acid derivatives (biotinand lipoic acid), and acts as an enzyme cofactor universal in three domains of the life. Although the biotin-requiring enzymes are rare proteins (in that mammals have only four such proteins whereas Escherichia coli has only a single biotinylated protein), they play critical roles in certain important reactions (like carboxylation, decarboxylation and trans-carboxylation) implicated into fatty acid synthesis, gluconeogenesis and amino acid degradation in both prokaryotes and eukaryotes. Right now, it is aware that most microorganisms (bacteria and fungi) and plants possess the ability to synthesize biotin, whereas mammals and birds cannot4. The earlier steps of biotin synthesis are involved in a modified type II fatty acid synthesis pathway in E. coli whereas the latter of biotin synthesis route refers to a highly-conserved four-step reactions catalyzed by BioF, BioA, BioD and BioB, respectively. In light that biotin is an energetically-expansive molecule in that generally ATP equivalents are estimated to be consumed via its paths of de novo synthesis for each biotin4, it seems reasonable that bacteria have developed diversified mechanisms to tightly monitor the level of biotin production in vivo. In addition to the paradigm E. coli BirA regulatory system that also retains the activity of biotin-protein ligase, at least two more regulatory machineries have been reported. Among them, one is the two-protein system of BirA coupled with BioR, the GntR-family transcription factor1, and the other denotes the two-protein system of the BirA linked to BioQ, a TetR family of transcription factor.
In fact, bacteria have evolved two different mechanisms to obtain the biotin cofactor for the metabolic requirement, one of which is de novo synthesis route, the other is a system of BioY transporter-mediated uptake. Unlike the human pathogen Brucella, a member of α-proteobacteria that encodes the above two systems for the availability of biotin11, it seems likely that the species of Streptococcus/Lactococcus family only have the BioY-based scavenging route and compensates the lack of the de novo biotin synthesis pathway. Among microorganisms, the paradigm enzyme with the biotin requirement refers to biotin carboxyl carrier protein (abbreviated as BCCP, i.e., the AccB subunit of acetyl-CoA carboxylase (ACC)) that catalyzes the first committed reaction for type II fatty acid synthesis pathway. Biotin protein ligase (BPL) is widespread in three domains of the life in that it transfers/attaches the biotin cofactor to the specific domain of the relevant subunits of key enzymes from the certain central metabolisms. Most of bacteria including E. colil and Bacillus only encode a single BPL to account for such kind of physiological requirement, while the pathogen Fracisella novicida developed an additional BPL to gain the competitive advantage in the infected host environment. In general, the BPL members are categorized into the following two groups (Group I and Group II) that can be easily distinguished by the presence of N-terminal DNA-binding domain that allow the BirA protein to bind the cognate genes (e.g., bio operon), and thereafter inhibit expression of biotin metabolism. Unlike the paradigm Group II BPL proteins, the E. coli birA gene product retaining the DNA-binding activity, the Group I BPL that lacks the N-terminal helix-turn-helix domain solely function as an enzyme responsible for protein biotinylation. In particular, the regulatory role of the Group II BPL depends on the participation of the physiological ligand/effector (biotinoyl-5′-AMP), the product of the first ligase half reaction for biotin utilization/protein biotinylation.
Resource: http://www.ncbi.nlm.nih.gov/
Resource: http://www.nutritionforest.com/
Biotin protein ligase is universal in three domains of life. The paradigm version of BPL is the Escherichia coli BirA that is also a repressor for the biotin biosynthesis pathway. Streptococcus suis, a leading bacterial agent for swine diseases, seems to be an increasingly-important opportunistic human pathogen. Unlike the scenario in E. coli, S. suis lacks the de novo biotin biosynthesis pathway. In contrast, it retains a bioY, a biotin transporter-encoding gene, indicating an alternative survival strategy for S. suis to scavenge biotin from its inhabiting niche. Here we report functional definition of S. suis birA homologue. The in vivo functions of the birA paralogue with only 23.6% identity to the counterpart of E. coli, was judged by its ability to complement the conditional lethal mutants of E. coli birA. The recombinant BirA protein of S. suis was overexpressed in E. coli, purified to homogeneity and verified with MS. Both cellulose TLC and MALDI-TOFF-MS assays demonstrated that the S. suis BirA protein catalyzed the biotinylation reaction of its acceptor biotin carboxyl carrier protein. EMSA assays confirmed binding of the bioY gene to the S. suis BirA. The data defined the first example of the bifunctional BirA ligase/repressor in Streptococcus.
Biotin (vitamin H) is one of two known sulfur-containing fatty acid derivatives (biotinand lipoic acid), and acts as an enzyme cofactor universal in three domains of the life. Although the biotin-requiring enzymes are rare proteins (in that mammals have only four such proteins whereas Escherichia coli has only a single biotinylated protein), they play critical roles in certain important reactions (like carboxylation, decarboxylation and trans-carboxylation) implicated into fatty acid synthesis, gluconeogenesis and amino acid degradation in both prokaryotes and eukaryotes. Right now, it is aware that most microorganisms (bacteria and fungi) and plants possess the ability to synthesize biotin, whereas mammals and birds cannot4. The earlier steps of biotin synthesis are involved in a modified type II fatty acid synthesis pathway in E. coli whereas the latter of biotin synthesis route refers to a highly-conserved four-step reactions catalyzed by BioF, BioA, BioD and BioB, respectively. In light that biotin is an energetically-expansive molecule in that generally ATP equivalents are estimated to be consumed via its paths of de novo synthesis for each biotin4, it seems reasonable that bacteria have developed diversified mechanisms to tightly monitor the level of biotin production in vivo. In addition to the paradigm E. coli BirA regulatory system that also retains the activity of biotin-protein ligase, at least two more regulatory machineries have been reported. Among them, one is the two-protein system of BirA coupled with BioR, the GntR-family transcription factor1, and the other denotes the two-protein system of the BirA linked to BioQ, a TetR family of transcription factor.
In fact, bacteria have evolved two different mechanisms to obtain the biotin cofactor for the metabolic requirement, one of which is de novo synthesis route, the other is a system of BioY transporter-mediated uptake. Unlike the human pathogen Brucella, a member of α-proteobacteria that encodes the above two systems for the availability of biotin11, it seems likely that the species of Streptococcus/Lactococcus family only have the BioY-based scavenging route and compensates the lack of the de novo biotin synthesis pathway. Among microorganisms, the paradigm enzyme with the biotin requirement refers to biotin carboxyl carrier protein (abbreviated as BCCP, i.e., the AccB subunit of acetyl-CoA carboxylase (ACC)) that catalyzes the first committed reaction for type II fatty acid synthesis pathway. Biotin protein ligase (BPL) is widespread in three domains of the life in that it transfers/attaches the biotin cofactor to the specific domain of the relevant subunits of key enzymes from the certain central metabolisms. Most of bacteria including E. colil and Bacillus only encode a single BPL to account for such kind of physiological requirement, while the pathogen Fracisella novicida developed an additional BPL to gain the competitive advantage in the infected host environment. In general, the BPL members are categorized into the following two groups (Group I and Group II) that can be easily distinguished by the presence of N-terminal DNA-binding domain that allow the BirA protein to bind the cognate genes (e.g., bio operon), and thereafter inhibit expression of biotin metabolism. Unlike the paradigm Group II BPL proteins, the E. coli birA gene product retaining the DNA-binding activity, the Group I BPL that lacks the N-terminal helix-turn-helix domain solely function as an enzyme responsible for protein biotinylation. In particular, the regulatory role of the Group II BPL depends on the participation of the physiological ligand/effector (biotinoyl-5′-AMP), the product of the first ligase half reaction for biotin utilization/protein biotinylation.
Resource: http://www.ncbi.nlm.nih.gov/
Resource: http://www.nutritionforest.com/
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