Limits...
The CacyBP/SIP protein is sumoylated in neuroblastoma NB2a cells.

Wasik U, Filipek A - Neurochem. Res. (2013)

Bottom Line: In this work we examined a post-translational modification of CacyBP/SIP which might have an effect on its function.By fractionation of NB2a cell extract we have found that, contrary to the majority of SUMO-modified proteins, sumoylated CacyBP/SIP is present in the cytoplasmic and not in the nuclear fraction.We have also established that lysine 16 is the residue which undergoes sumoylation in the CacyBP/SIP protein.

View Article: PubMed Central - PubMed

ABSTRACT
The Calcyclin binding protein and Siah-1 interacting protein (CacyBP/SIP) protein is highly expressed in mammalian brain as well as in neuroblastoma NB2a cells and pheochromocytoma PC12 cells. This protein interacts with several targets such as cytoskeletal proteins or ERK1/2 kinase and seems to be involved in many cellular processes. In this work we examined a post-translational modification of CacyBP/SIP which might have an effect on its function. Since theoretical analysis of the amino acid sequence of CacyBP/SIP indicated several lysine residues which could potentially be sumoylated we checked experimentally whether this protein might be modified by SUMO attachment. We have shown that indeed CacyBP/SIP bound the E2 SUMO ligase, Ubc9, in neuroblastoma NB2a cell extract and was sumoylated in these cells. By fractionation of NB2a cell extract we have found that, contrary to the majority of SUMO-modified proteins, sumoylated CacyBP/SIP is present in the cytoplasmic and not in the nuclear fraction. We have also established that lysine 16 is the residue which undergoes sumoylation in the CacyBP/SIP protein.

Show MeSH

Related in: MedlinePlus

Model of the CacyBP/SIP N-terminal fragment (NTF) with bound SUMO1. The NTF structure of mouse CacyBP/SIP was taken from the Protein Data Bank (PDB; id:1YSM) as the first model of the 20 NMR structures [27]. The structure of human SUMO1, which shares 100 % amino acid sequence identity with the mouse ortholog, was extracted from a crystal structure of the human SUMO E1 complex (PDB; id:3KYC) [28]. Both proteins were bonded via residues K16 from NTF of CacyBP/SIP and G97 from the SUMO1 C-terminus. The K16 residue of CacyBP/SIP (red) and SUMO1 molecule are shown in semitransparent surfaces (Color figure online)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3824344&req=5

Fig4: Model of the CacyBP/SIP N-terminal fragment (NTF) with bound SUMO1. The NTF structure of mouse CacyBP/SIP was taken from the Protein Data Bank (PDB; id:1YSM) as the first model of the 20 NMR structures [27]. The structure of human SUMO1, which shares 100 % amino acid sequence identity with the mouse ortholog, was extracted from a crystal structure of the human SUMO E1 complex (PDB; id:3KYC) [28]. Both proteins were bonded via residues K16 from NTF of CacyBP/SIP and G97 from the SUMO1 C-terminus. The K16 residue of CacyBP/SIP (red) and SUMO1 molecule are shown in semitransparent surfaces (Color figure online)

Mentions: To explain the lack of effect of CacyBP/SIP sumoylation on its functions we have analyzed the structure of the modified protein. For that we have employed a molecular modeling method using the available structures of CacyBP/SIP N-terminus and of SUMO1. As it can be seen in Fig. 4, the N-terminal part of CacyBP/SIP forms two anti-parallel helices (47 amino acids). The K16 residue is located on the side of the helix not facing the other helix therefore the attachment of a moiety, even one as large as SUMO1, should not significantly affect the structure of CacyBP/SIP. The exposed location of the SUMO moiety on CacyBP/SIP makes it easily accessible for other proteins. Interestingly, sumoylation, similarly to some other posttranslational modifications, for instance acetylation, is regarded as a modification that may help to assemble large protein complexes since the SUMO moiety can be recognized by a SUMO interacting motif (SIM) present in other proteins [25]. Thus, CacyBP/SIP sumoylation may facilitate its interaction with other protein partners. This feature is important since CacyBP/SIP was postulated to be a component of multiprotein complexes [26].Fig. 4


The CacyBP/SIP protein is sumoylated in neuroblastoma NB2a cells.

Wasik U, Filipek A - Neurochem. Res. (2013)

Model of the CacyBP/SIP N-terminal fragment (NTF) with bound SUMO1. The NTF structure of mouse CacyBP/SIP was taken from the Protein Data Bank (PDB; id:1YSM) as the first model of the 20 NMR structures [27]. The structure of human SUMO1, which shares 100 % amino acid sequence identity with the mouse ortholog, was extracted from a crystal structure of the human SUMO E1 complex (PDB; id:3KYC) [28]. Both proteins were bonded via residues K16 from NTF of CacyBP/SIP and G97 from the SUMO1 C-terminus. The K16 residue of CacyBP/SIP (red) and SUMO1 molecule are shown in semitransparent surfaces (Color figure online)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3824344&req=5

Fig4: Model of the CacyBP/SIP N-terminal fragment (NTF) with bound SUMO1. The NTF structure of mouse CacyBP/SIP was taken from the Protein Data Bank (PDB; id:1YSM) as the first model of the 20 NMR structures [27]. The structure of human SUMO1, which shares 100 % amino acid sequence identity with the mouse ortholog, was extracted from a crystal structure of the human SUMO E1 complex (PDB; id:3KYC) [28]. Both proteins were bonded via residues K16 from NTF of CacyBP/SIP and G97 from the SUMO1 C-terminus. The K16 residue of CacyBP/SIP (red) and SUMO1 molecule are shown in semitransparent surfaces (Color figure online)
Mentions: To explain the lack of effect of CacyBP/SIP sumoylation on its functions we have analyzed the structure of the modified protein. For that we have employed a molecular modeling method using the available structures of CacyBP/SIP N-terminus and of SUMO1. As it can be seen in Fig. 4, the N-terminal part of CacyBP/SIP forms two anti-parallel helices (47 amino acids). The K16 residue is located on the side of the helix not facing the other helix therefore the attachment of a moiety, even one as large as SUMO1, should not significantly affect the structure of CacyBP/SIP. The exposed location of the SUMO moiety on CacyBP/SIP makes it easily accessible for other proteins. Interestingly, sumoylation, similarly to some other posttranslational modifications, for instance acetylation, is regarded as a modification that may help to assemble large protein complexes since the SUMO moiety can be recognized by a SUMO interacting motif (SIM) present in other proteins [25]. Thus, CacyBP/SIP sumoylation may facilitate its interaction with other protein partners. This feature is important since CacyBP/SIP was postulated to be a component of multiprotein complexes [26].Fig. 4

Bottom Line: In this work we examined a post-translational modification of CacyBP/SIP which might have an effect on its function.By fractionation of NB2a cell extract we have found that, contrary to the majority of SUMO-modified proteins, sumoylated CacyBP/SIP is present in the cytoplasmic and not in the nuclear fraction.We have also established that lysine 16 is the residue which undergoes sumoylation in the CacyBP/SIP protein.

View Article: PubMed Central - PubMed

ABSTRACT
The Calcyclin binding protein and Siah-1 interacting protein (CacyBP/SIP) protein is highly expressed in mammalian brain as well as in neuroblastoma NB2a cells and pheochromocytoma PC12 cells. This protein interacts with several targets such as cytoskeletal proteins or ERK1/2 kinase and seems to be involved in many cellular processes. In this work we examined a post-translational modification of CacyBP/SIP which might have an effect on its function. Since theoretical analysis of the amino acid sequence of CacyBP/SIP indicated several lysine residues which could potentially be sumoylated we checked experimentally whether this protein might be modified by SUMO attachment. We have shown that indeed CacyBP/SIP bound the E2 SUMO ligase, Ubc9, in neuroblastoma NB2a cell extract and was sumoylated in these cells. By fractionation of NB2a cell extract we have found that, contrary to the majority of SUMO-modified proteins, sumoylated CacyBP/SIP is present in the cytoplasmic and not in the nuclear fraction. We have also established that lysine 16 is the residue which undergoes sumoylation in the CacyBP/SIP protein.

Show MeSH
Related in: MedlinePlus