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. 2007 Jul 10;581(17):3164-70.
doi: 10.1016/j.febslet.2007.05.082. Epub 2007 Jun 12.

The maximal size of protein to diffuse through the nuclear pore is larger than 60kDa

Ruiwen Wang  1 , Michael G Brattain
Affiliations

The maximal size of protein to diffuse through the nuclear pore is larger than 60kDa

Ruiwen Wang et al. FEBS Lett. .

Abstract

It has generally been believed that the diffusion limit set by the nuclear pore for protein is 60kDa. We here studied the cellular localization of several artificial proteins and found that the diffusion limit set by the nuclear pore is not as small as previously thought. The results indicate that the maximal size of protein to diffuse through the nuclear pore complex could be quite larger than 60kDa, thus greatly extending the diffusion limit that the nuclear pore can accommodate.

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Figures

Fig. 1
Fig. 1
Mapping of the largest chimeric GFP protein that is allowed to diffuse through the nuclear pore. (A) Schematic representation of chimeric GFP oligomer proteins. (B) Western blot showing expression and migration of chimeric GFP oligomer proteins. Lysates were collected at 16 hours after transfection with the indicated constructs in HEK293 cells and the molecular weights of the transfected proteins were determined by Western blot with mouse antibody against GFP. (C) The cellular localization of the chimeric GFP proteins was visualized at 8 hours after transient transfection with indicated constructs in HeLa cells.
Fig. 2
Fig. 2
Mapping of the maximal size of protein based on ERK2 that is allowed to diffuse through the nuclear pore. (A) The cellular localization of the GFP5 fusion proteins was visualized at 8 hours after transient transfection with indicated constructs in HeLa cells. (B) Schematic representation of chimeric ERK2 fusion proteins. (C) Western blot showing expression and migration of chimeric ERK2 fusion proteins. Lysates were collected at 16 hours after transfection with the indicated constructs and the molecular weights of the transfected proteins were determined by Western blot with mouse antibody against Myc tag. (D) The cellular localization of the chimeric fusion proteins based on Myc-ERK2 was visualized at 8 hours after transient transfection with indicated constructs.
Fig. 3
Fig. 3
Mapping of the maximal size of protein based on PDK1 that is allowed to diffuse through the nuclear pore. (A) The cellular localization of the wt or mutant PDK1 protein was visualized at 8 hours after transient transfection with indicated constructs. (B) Schematic representation of chimeric PDK1 fusion proteins. (C) Western blots showing expression and migration of chimeric PDK1 fusion proteins. Lysates were collected at 16 hours after transfection with the indicated constructs and the molecular weights of the transfected proteins were determined by Western blot with mouse antibody against Myc tag. (D) The cellular localization of the chimeric fusion proteins based on Myc-PDK1ΔNES was visualized at 8 hours after transient transfection with indicated constructs. a: Myc-PDK1ΔNES+100aa; b: Myc-PDK1ΔNES+135aa; c: Myc-PDK1ΔNES+200aa; d: Myc-PDK1ΔNES+300aa.
Fig. 3
Fig. 3
Mapping of the maximal size of protein based on PDK1 that is allowed to diffuse through the nuclear pore. (A) The cellular localization of the wt or mutant PDK1 protein was visualized at 8 hours after transient transfection with indicated constructs. (B) Schematic representation of chimeric PDK1 fusion proteins. (C) Western blots showing expression and migration of chimeric PDK1 fusion proteins. Lysates were collected at 16 hours after transfection with the indicated constructs and the molecular weights of the transfected proteins were determined by Western blot with mouse antibody against Myc tag. (D) The cellular localization of the chimeric fusion proteins based on Myc-PDK1ΔNES was visualized at 8 hours after transient transfection with indicated constructs. a: Myc-PDK1ΔNES+100aa; b: Myc-PDK1ΔNES+135aa; c: Myc-PDK1ΔNES+200aa; d: Myc-PDK1ΔNES+300aa.
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