For the first time, the new method details the assembly process of the nuclear hole complex!

Cells produce a large number of protein complexes, each of which is made up of many individual proteins.
protein complexes, such as ucose, regulate almost all life-sustaining biological functions in cells.
have succeeded in determining the structure of many of these complexes, but so far there has been little research on how individual proteins are assembled and change over time.
, traditional methods have proven insufficient to study the exact processes of these reactions in cells, especially when it comes to large protein complexes.
methods make it possible to track dynamic changes in protein complex assembly, even very large protein complexes, with high time resolution.
inspired by metabolic analysis, the researchers called their new method KARMA (kinetic analysis of incorporation rates in macromolecular assembly, a dynamic analysis of the rate of mixing in polymer assembly), based on methods that study metabolic processes.
scientists who study metabolism have long used radioactive carbon in their work, for example, to label glucose molecules, which are then absorbed and metabolized by cells.
this radioactive marker allows people to track the location and point at which glucose molecules or their metabolites appear.
wes explained, "This type of study inspired us to apply similar principles to explore the reactions that occur during the assembly of protein complexes.
their method, the researchers studied using labeled amino acids, the basic components of the protein that contain heavier carbon and nitrogen isotopes.
in yeast cell cultures, they replace lighter amino acids with heavier amino acids.
yeasts use these heavier amino acids in protein synthesis, resulting in changes in the molecular weight of all newly produced proteins.
-scale time scale for the assembly of the
complex In order to isolate the protein complex, the researchers took yeast cells from the culture every once in a while and used mass spectrometers to measure the small weight difference between molecules with heavier amino acids and molecules without heavier amino acids.
indicates the age of the protein in the protein complex.
, the older the protein, the earlier it is integrated into the complex.
these age differences, they applied dynamic state models to eventually reconstruct the exact assembly order of specific protein complexes.
as a case study to test this new approach, Weiss and his team chose a nuclear hole complex in yeast cells.
structure has about 500 to 1,000 components and consists of about 30 different proteins, each with multiple copies, making it one of the largest protein complexes known.
KARMA, the researchers were able to obtain detailed maps of which components were integrated into the structure and when.
of their findings is the hierarchical principle: individual proteins form sub-fundamentals in a very short period of time and then assemble them from the center to the periphery in a specific order.
for the first time, we have confirmed that some proteins are integrated very quickly during the assembly of the nucleo-hole complex, while others are integrated after about an hour, " says Weiss, a durable internal stent.
it's been an incredibly long time.
" yeast cells divide every 90 minutes, meaning it takes almost a generation to assemble this important nuclear compound.
it is not clear exactly why the assembly of the new nucleus takes so long to be related to the yeast's reproductive cycle.
the researchers also found that once the core holes were assembled, the various parts of the core hole complex in the internal bracket were highly stable and durable, with almost no components replaced during their life cycle.
, proteins on the periphery of the nucleo-hole complex are often replaced.
defective nucleokers promote disease The nucleokers are one of the most important protein complexes in cells because they are responsible for the exchange of matter and molecules between the nucleosytes and cytons.
, for example, transport messenger RNA from the nucleation to cell machines outside the nucleo, which need these molecules as a basis for making new proteins.
addition, nucleoker holes play a direct and indirect role in human diseases.
, changes in the nucleosis and its proteins can affect the production of neurodegenerative diseases such as leukemia, diabetes or Alzheimer's disease.
, "But generally speaking, the cause of these disease patterns is not clear."
," he explains, karma may help to gain a deeper understanding of such issues in the future.
, a multipurpose platform, said, "Although we used KARMA in this study for only one protein complex, we are excited about its future application."
approach will now allow us to decipher the sequence in which a series of biological processes occur.
, for example, their techniques can be used to study molecular events that occur during the infection cycle of a virus, such as SARS-CoV-2, and may help find new candidates to disrupt the infection cycle.
" this new method can also be applied to biological molecules other than proteins, such as RNA or lipids.
: Evgeny Onischenko et al. Maturation Kinetics of a Multiprotein Complex Revealed by Metabolic Labeling. Cell, 2020, doi:10.1016/j.cell.2020.11.001.