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The large subunit of Leishmania topoisomerase I functions as the ‘molecular steer’ in type IB topoisomerase
Author(s) -
BoseDasgupta Somdeb,
Ganguly Agneyo,
Das Benu Brata,
Roy Amit,
Khalkho Neeta V. M.,
Majumder Hemanta K.
Publication year - 2008
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2007.06002.x
Subject(s) - biology , topoisomerase , protein subunit , dna , minicircle , biochemistry , cleavage (geology) , microbiology and biotechnology , gene , paleontology , fracture (geology)
Summary Kinetoplastid topoisomerase IB is an unusual bisubunit enzyme where reconstitution of the large (LdTOPIL or L) and small (LdTOPIS or S) subunits shows functional activity. It is yet to be deciphered whether one subunit or both navigate the heterodimer to its cellular DNA targets. Tethering a specific DNA‐binding protein to topoisomerase I alters its site specificity. The chimeric constructs UMSBP–LdTOPIL/S or U–L/S (fusion of UMSBP to the N‐terminus of L and reconstituted with S) and LdTOPIL/UMSBP–LdTOPIS or L/U–S (fusion of UMSBP to the N‐terminus of S and reconstituted with L) exhibit relaxation activity. Only U–L/S shows altered site specificity and enhanced DNA‐binding affinity for the universal minicircle sequence (UMS) containing substrate. This proves that L alone serves as the ‘molecular steer’ for this heterodimer. Reconstituted U–L/S also induces cleavage close to UMS and causes minicircle linearization. The differential properties of the reconstituted chimeras U–L/S and L/U–S reveal the structural and functional asymmetry between the heterodimer. Therefore this study helps in a better understanding of the mechanistic details underlying topoisomerization by this bi‐subunit enzyme.