Size of Human Lens β-Crystallin Aggregates Are Distinguished by N-terminal Truncation of βB1

Abstract

The aggregates formed by the interactions of the human lens β-crystallins have been particularly difficult to characterize because the β-crystallins comprise several proteins of similar structure and molecular weight and because their sequences were not known until recently. Previously, it could not be ascertained whether the species of various acidities were different proteins or modifications of the same proteins. The recent determination of the sequences permits calculation of molecular weights and unambiguous identification of the various β-crystallins and their modified forms by mass spectrometry. In this investigation, the components of the three sizes of β-crystallin aggregates, β1 (∼150,000), β2 (∼92,000), and β3 (∼46,000), were determined. The principal differences among the different β-crystallin aggregates was the presence of βA4 in β1 and β2, but not β3, and the length of the N-terminal extension of βB1. The size of the β-crystallin aggregate correlated with the length of the N-terminal extension of βB1, indicating that the flexible N terminus of βB1 is critical to the formation of higher molecular weight aggregates of β-crystallins. Separation of the components by ion exchange under non-denaturing conditions showed that βB2 occurs as homo-dimers and homo-tetramers as well as contributing to hetero-oligomers. Other β-crystallins were present only as hetero-oligomers. The aggregates formed by the interactions of the human lens β-crystallins have been particularly difficult to characterize because the β-crystallins comprise several proteins of similar structure and molecular weight and because their sequences were not known until recently. Previously, it could not be ascertained whether the species of various acidities were different proteins or modifications of the same proteins. The recent determination of the sequences permits calculation of molecular weights and unambiguous identification of the various β-crystallins and their modified forms by mass spectrometry. In this investigation, the components of the three sizes of β-crystallin aggregates, β1 (∼150,000), β2 (∼92,000), and β3 (∼46,000), were determined. The principal differences among the different β-crystallin aggregates was the presence of βA4 in β1 and β2, but not β3, and the length of the N-terminal extension of βB1. The size of the β-crystallin aggregate correlated with the length of the N-terminal extension of βB1, indicating that the flexible N terminus of βB1 is critical to the formation of higher molecular weight aggregates of β-crystallins. Separation of the components by ion exchange under non-denaturing conditions showed that βB2 occurs as homo-dimers and homo-tetramers as well as contributing to hetero-oligomers. Other β-crystallins were present only as hetero-oligomers.