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Defective folding of mutant p16(INK4) proteins encoded by tumor-derived alleles
1Department of Biochemistry, University of Connecticut Health Center, Farmington, Connecticut 06032, USA. peng@sun.uchc.edu
Abstract:
p16(INK4) is a specific cyclin D-dependent kinase inhibitor and a multiple tumor suppressor. Inactivation of p16 is frequent in both primary tumors and tumor-derived cell lines. We describe here the conformational properties and oligomerization state of seven mutant p16 proteins; all of them are deficient in function. Four of the seven proteins show significantly disrupted secondary structure and backbone folding. The other three adopt partially folded, molten globule-like conformations. These proteins have near-native levels of secondary structure, but lack the ability to undergo a cooperative thermal transition and are substantially less resistant to proteolysis than is wild type p16. At low concentrations, two of the seven proteins are monomers, three exhibit an apparent molecular weight between the value of a monomer and a dimer, and the other two aggregate significantly. Our results strongly suggest that defective protein folding and/or aggregation is a common mechanism for inactivation of p16.
Insights
Mutant p16 proteins, crucial tumor suppressors, often lose function due to misfolding or aggregation. This study reveals how altered protein structures and oligomerization states contribute to p16INK4 inactivation in cancer.
Area of Science:
- Molecular Biology
- Biochemistry
- Cancer Research
Background:
- p16(INK4) is a critical tumor suppressor and cyclin D-dependent kinase inhibitor.
- Inactivation of p16 is a common event in various cancers.
Purpose of the Study:
- To investigate the conformational properties and oligomerization of functionally deficient p16 mutants.
- To elucidate the role of protein folding and aggregation in p16 inactivation.
Main Methods:
- Analysis of secondary structure and backbone folding of seven mutant p16 proteins.
- Assessment of thermal stability and proteolysis resistance.
- Determination of oligomerization states (monomer, dimer, aggregation) using molecular weight estimations.
Main Results:
- Four mutants displayed disrupted secondary structure and folding.
- Three mutants adopted molten globule-like conformations with altered stability and proteolysis resistance.
- Oligomerization varied, with monomers, intermediate species, and significant aggregation observed.
Conclusions:
- Defective protein folding is a significant mechanism for p16(INK4) inactivation.
- Protein aggregation also contributes to the loss of p16 tumor suppressor function.
- Understanding these mechanisms can inform cancer therapy development.