Brain-specific metallothionein-3 has higher metal-binding capacity than ubiquitous metallothioneins and binds metals noncooperatively

Biochemistry. 2002 May 14;41(19):6158-63. doi: 10.1021/bi025664v.

Abstract

Zinc metabolism in the cells is largely regulated by ubiquitous small proteins, metallothioneins (MT). Metallothionein-3 is specifically expressed in the brain and is down regulated in Alzheimer's disease. We demonstrate by mass spectrometry that MT-3, in contrast to common MTs, binds Zn(2+) and Cd(2+) in a noncooperative manner and can also bind higher stoichiometries of metals than seven. MT-3 reconstituted with seven metals exists in a dynamic equilibrium of different metalloforms, where the prevalent metalloform is Me(7)MT-3, but metalloforms with 6, 8, and even 9 metals are also present. The results from pH and stability studies demonstrate that the heterogeneity of metalloforms originates from the N-terminal beta-cluster, whereas the C-terminal alpha-cluster of MT-3 binds four metal ions such as that of common MTs. Experiments with EDTA demonstrate that the beta-cluster of ZnMT-3 has a higher metal transfer potential than the beta-cluster of Zn(7)MT-2. Moreover, ZnMT-3 loses metals during ultrafiltration. MT-3, reconstituted with an excess of Zn(2+) or Cd(2+), exists as a dynamic mixture of metalloforms with higher than 7 metal stoichiometries (8-11). Such forms of ZnMT-3 are unstable and decompose partly already during a rapid gel filtration, whereas CdMT-3 forms are more stable. Extra metal ions may bind to the beta-cluster region as well as to the carboxylates of MT-3. The specific metal-binding properties of MT-3 could be functionally implemented for buffering of fluctuating concentrations of zinc in zincergic neurons and for transfer of zinc to synaptic vesicles.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alzheimer Disease / metabolism
  • Animals
  • Brain / metabolism*
  • Cadmium / metabolism
  • Humans
  • In Vitro Techniques
  • Isoenzymes / chemistry
  • Isoenzymes / metabolism
  • Kinetics
  • Liver / metabolism
  • Metallothionein 3
  • Metals / metabolism*
  • Nerve Tissue Proteins / chemistry*
  • Nerve Tissue Proteins / metabolism*
  • Rabbits
  • Spectrometry, Mass, Electrospray Ionization
  • Zinc / metabolism

Substances

  • Isoenzymes
  • Metallothionein 3
  • Metals
  • Nerve Tissue Proteins
  • Cadmium
  • Zinc