The Continental Shelf Beyond 200 Nautical Miles

As policymakers, academia, and the media have paid increased attention to the Arctic region, there is more evidence of a certain lack of knowledge concerning the applicable international law. The United Nations Convention on the Law of the Sea of December 10, 1982--adopted in 1982 and in force since November 16, 1994--provides both a legal framework within which all activities in oceans and seas must be carried out and, as far as the seabed of the Arctic Ocean international law is concerned, answers to questions related to its legal status and applicable regulations. If a coastal State wishes to delineate its continental shelf beyond 200 nautical miles from the baselines from which the breadth of the territorial sea is measured, it has to submit relevant data and information to the Commission on the Limits of the Continental Shelf, an expert body established under the Convention. The Commission issues recommendations, and the limits based on the recommendations of that Commission are final and binding. In the Arctic region, only two coastal States so far have made submissions to the Commission--the Russian Federation and Norway. The Commission issued recommendations to both; in the case of the Central Arctic Ocean, it recommended that the Russian Federation make a revised submission. Due to the fact that the other three coastal States of the Arctic Ocean--Canada, Denmark, and the United States--have yet to make their submissions (the United States is still not party to the Convention), and taking into account the workload of the Commission, the delineation of the continental shelf beyond 200 nautical miles and related delimitation of maritime boundaries between States will take many years to finalize

Fluorescence Polarization and Fluctuation Analysis Monitors Subunit Proximity, Stoichiometry, and Protein Complex Hydrodynamics

Förster resonance energy transfer (FRET) microscopy is frequently used to study protein interactions and conformational changes in living cells. The utility of FRET is limited by false positive and negative signals. To overcome these limitations we have developed Fluorescence Polarization and Fluctuation Analysis (FPFA), a hybrid single-molecule based method combining time-resolved fluorescence anisotropy (homo-FRET) and fluorescence correlation spectroscopy. Using FPFA, homo-FRET (a 1–10 nm proximity gauge), brightness (a measure of the number of fluorescent subunits in a complex), and correlation time (an attribute sensitive to the mass and shape of a protein complex) can be simultaneously measured. These measurements together rigorously constrain the interpretation of FRET signals. Venus based control-constructs were used to validate FPFA. The utility of FPFA was demonstrated by measuring in living cells the number of subunits in the α-isoform of Venus-tagged calcium-calmodulin dependent protein kinase-II (CaMKIIα) holoenzyme. Brightness analysis revealed that the holoenzyme has, on average, 11.9±1.2 subunit, but values ranged from 10–14 in individual cells. Homo-FRET analysis simultaneously detected that catalytic domains were arranged as dimers in the dodecameric holoenzyme, and this paired organization was confirmed by quantitative hetero-FRET analysis. In freshly prepared cell homogenates FPFA detected only 10.2±1.3 subunits in the holoenzyme with values ranging from 9–12. Despite the reduction in subunit number, catalytic domains were still arranged as pairs in homogenates. Thus, FPFA suggests that while the absolute number of subunits in an auto-inhibited holoenzyme might vary from cell to cell, the organization of catalytic domains into pairs is preserved