Role of small loops in DNA melting

Short melted regions less than 100 base pairs (bp) in length are rarely fou nd in the differential melting curves (DMC) of natural DNAs. Therefore, it is supposed that their characteristics do not affect DNA melting behavior. However. in our previous study, a strong influence of the form of the entro py factor of small loops on melting of cross-linked DNAs was established (D . Y, Lando, A. S. Fridman et al., Journal of Biomolecular Structure and Dyn amics, 1997, Vol. 15, pp. 141-150; Journal of Biomolecular Structure and Dy namics, 1998, Vol. 16, pp. 59-67). Quite different dependencies of the melt ing temperature on the relative concentration of interstrand cross-links we re obtained for the loop entropy factors given by the Fixman-Freire (Jacobs on-Stockmayer) and Wartell-Benight relations. In the present study, the inf luence of the entropy factor of small loops on the melting of natural DNAs, cross-linked DNAs and periodical double-stranded polynucleotides is compar ed using computer simulation. A fast combined computational method for calc ulating DNA melting curves was developed for this investigation, It allows us to assign an arbitrary dependence of the loop entropy factor on the leng th of melted regions for the terms corresponding to small loops (less than tau bp in length). These terms are calculated using Poland's approach. The Fixman-Freire approach is used for long loops. Our calculations have shown that the temperature dependence of the average length of interior melted re gions (loops) has a maximum at T approximate to T-m (T-m is the DNA melting temperature) in contrast to the dependence of the total average length of melted regions, which increases almost monotonously. Computer modeling demo nstrates that prohibition of formation of loops less than tau base pairs in length does nor markedly change the DMC for tau < 150 bp. However, the sam e prohibition strongly affects the average length of internal melted region s for much smaller <tau>'s. The effect is already noticeable for tau = 1 bp and increases with T. A tenfold increase in the entropy factor of all loop s with length less than tau bp causes a noticeable alteration of the DMC fo r tau greater than or equal to 30 bp. It is shown that DMCs are identical f or the Wartell-Benight and for the Fixman-Freire (Jacobson-Stockmayer) form of the loop entropy factor However, for low degree of denaturation, the av erage length of internal melted regions is 40% lower for the Wartell-Benigh t form due to the fluctuational opening of short AT-rich regions less than 10 bp in length. The same calculations carried out for periodical polynucle otides demonstrate a much stronger difference in melting behavior for diffe rent forms of entropy factors of short loops. The strongest difference occu rs if the length of stable GC-rich and unstable AT-rich stretches is equal to 30 bp. However, the comparison carried out in this work demonstrates tha t the entropy factor of short loops influences melting behavior of cross-li nked DNA much stronger than of unmodified DNA with random or periodical seq uences. (C) 2001 John Wiley & Sons. Inc. Biopolymers 58: 374-389, 2001.