title: The amino-terminus of nitric oxide sensitive guanylyl cyclase α₁ does not affect dimerization but influences subcellular localization.
abstract: BACKGROUND
  Nitric oxide sensitive guanylyl cyclase (NOsGC) is a heterodimeric enzyme formed by an α- and a β₁-subunit. A splice variant (C-α₁) of the α₁-subunit, lacking at least the first 236 amino acids has been described by Sharina et al. 2008 and has been shown to be expressed in differentiating human embryonic cells. Wagner et al. 2005 have shown that the amino acids 61-128 of the α₁-subunit are mandatory for quantitative heterodimerization implying that the C-α₁-splice variant should lose its capacity to dimerize quantitatively.
  
  
  RESULTS
  In the current study we demonstrate preserved quantitative dimerization of the C-α₁-splice by co-purification with the β₁-subunit. In addition we used fluorescence resonance energy transfer (FRET) based on fluorescence lifetime imaging (FLIM) using fusion proteins of the β₁-subunit and the α₁-subunit or the C-α₁ variant with ECFP or EYFP. Analysis of the respective combinations in HEK-293 cells showed that the fluorescence lifetime was significantly shorter (≈0.3 ns) for α₁/β₁ and C-α₁/β₁ than the negative control. In addition we show that lack of the amino-terminus in the α₁ splice variant directs it to a more oxidized subcellular compartment.
  
  
  CONCLUSIONS
  We conclude that the amino-terminus of the α₁-subunit is dispensable for dimerization in-vivo and ex-vivo, but influences the subcellular trafficking.
authors: Department of Pharmacology, Toxicology and Clinical Pharmacy, University of Brunswick - Institute of Technology, Brunswick, Germany.
authors: Jan R
authors: JR
authors: Kraehling
authors: Mareike
authors: M
authors: Busker
authors: Tobias
authors: T
authors: Haase
authors: Nadine
authors: N
authors: Haase
authors: Markus
authors: M
authors: Koglin
authors: Monika
authors: M
authors: Linnenbaum
authors: Soenke
authors: S
authors: Behrends
mesh_terms: D002460:Cell Line; D031541:Fluorescence Resonance Energy Transfer; D006162:Guanylate Cyclase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D021122:Protein Subunits; D006162:Guanylate Cyclase