Barley contains two different isoforms of flavin-containing polyamine oxidase (BPAO1 and BPAO2). We havepreviously demonstrated that BPAO2 is a symplastic protein in barley leaves. On the contrary, maizepolyamine oxidase (MPAO), the best characterized member of this enzyme class, is apoplastic. Comparison ofthe derived amino-acid sequences of BPAO2 and MPAO has revealed that both precursor proteins include acleavable N-terminal signal peptide of 25 amino acid residues, but the barley enzyme shows an extraC-terminal extension of eight amino acids. By means of MPAO engineering with BPAO2 C-terminal tail (MPAOT)and exploiting transient expression in Nicotiana tabacum protoplasts, we demonstrate that thisoligopeptide is a signal for protein sorting to the plant vacuole. The vacuolar sorting of MPAO-T wassaturable. Specific mutations of the C-terminal tail were constructed to determine which amino acid residuesof this novel propeptide affect proper protein sorting. No consensus sequence or common structuraldeterminant is required for the intracellular retention of the MPAO-T protein, but a gradual lowering of theefficiency was observed as a result of progressive deletion of the C-terminus.
A novel C-terminal sequence from barley polyamine oxidase is a vacuolar sorting signal.
Vitale A;
2004
Abstract
Barley contains two different isoforms of flavin-containing polyamine oxidase (BPAO1 and BPAO2). We havepreviously demonstrated that BPAO2 is a symplastic protein in barley leaves. On the contrary, maizepolyamine oxidase (MPAO), the best characterized member of this enzyme class, is apoplastic. Comparison ofthe derived amino-acid sequences of BPAO2 and MPAO has revealed that both precursor proteins include acleavable N-terminal signal peptide of 25 amino acid residues, but the barley enzyme shows an extraC-terminal extension of eight amino acids. By means of MPAO engineering with BPAO2 C-terminal tail (MPAOT)and exploiting transient expression in Nicotiana tabacum protoplasts, we demonstrate that thisoligopeptide is a signal for protein sorting to the plant vacuole. The vacuolar sorting of MPAO-T wassaturable. Specific mutations of the C-terminal tail were constructed to determine which amino acid residuesof this novel propeptide affect proper protein sorting. No consensus sequence or common structuraldeterminant is required for the intracellular retention of the MPAO-T protein, but a gradual lowering of theefficiency was observed as a result of progressive deletion of the C-terminus.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
