Effects of tls parameters in macromolecular refinement
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Effects of TLS parameters in Macromolecular Refinement. Martyn Winn Daresbury Laboratory, U.K. IUCr99 08/08/99. Overview. Background to the use of TLS tensors. Details of TLS refinement. Implementation in REFMAC: examples. Contributions to atomic U.

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Effects of TLS parameters in Macromolecular Refinement

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Effects of tls parameters in macromolecular refinement

Effects of TLS parameters in Macromolecular Refinement

  • Martyn Winn

  • Daresbury Laboratory, U.K.

  • IUCr99 08/08/99


Overview

Overview

  • Background to the use of TLS tensors.

  • Details of TLS refinement.

  • Implementation in REFMAC: examples


Contributions to atomic u

Contributions to atomic U

U = Ucrystal + UTLS + Uinternal + Uatom

Ucrystal : overall anisotropic scale factor w.r.t. crystal axes.

UTLS : rigid body displacements e.g. of a.s.u., molecules, domains, secondary structure elements,

aromatic rings of side groups, etc.

Uinternal : internal displacements of molecules, e.g. normal modes of vibration, torsions, etc.

Uatom : anisotropy of individual atoms


Tls small molecules

TLS: small molecules

  • D.W.J.Cruikshank (1956) - TL analysis

  • G.S.Pawley (1964, 1966) - TL refinement

  • V.Schomaker & K.N.Trueblood (1968) - introduction of S in analysis of ADPs

  • J.D.Dunitz & D.N.J.White (1973) - inclusion of internal torsional motion of “attached rigid group”


Tls macromolecules

TLS: macromolecules

  • S.R.Holbrook et al (1985) - duplex DNA dodecanucleatide, 1.9Å, 70 groups (phosphate, ribose, base), CORELS

  • B.Howlin et al (1989) - bovine Ribonuclease A, 1.45Å, RESTRAIN

  • G.W.Harris et al (1992) - papain, 1.6Å , RESTRAIN

  • Sali et al (1992) - endothiapepsin complex, 1.8Å, RESTRAIN


Rigid body motion

Rigid body motion

  • Linearise general displacement u of atom (mean position r) in rigid body: u = t + D.r  t +  x r

  • Corresponding dyad: uu = tt + t x r - r x t - r x  x r

  • Average over dynamic motion and static disorder gives atomic ADP: U  <uu> = T + ST x r - r x S - r x L x r

  • T,L andS describe mean square translation, libration and their correlation of rigid body.


Tls in refinement

TLS in refinement

  • Need to specify TLS groups for molecule of interest.

  • 6 + 6 + 8 = 20 parameters per group (trace of S is undetermined).

  • T and S (but not L) origin-dependent. S is symmetric if origin is Centre of Reaction.

  • Gradients of residual w.r.t. TLS parameters follow from gradients w.r.t. U’s via chain rule.


Effects of tls parameters in macromolecular refinement

NCS

  • REFMAC applies restraints to B and U values of NCS-related molecules.

  • But different molecules in a.s.u. may have different overall thermal parameters.

  • Refine independent overall TLS tensors for each molecule before refining restrained individual parameters.


Choice of tls groups

Choice of TLS groups

  • Choose TLS groups using:

    • chemical knowledge, e.g. aromatic side groups of amino acids, domains of macromolecules

    • fit to ADPs of test structure, e.g. Holbrook & Kim (1984) compared 7 rigid body models of CMP and used best as basis for partitioning other nucleic acids.

    • dynamic domains identified from similar structures, e.g. from apo and holo forms of alcohol dehydrogenase


Implementation in refmac

Implementation in REFMAC

  • Refine TLS parameters against ML residual, using previously refined atomic coordinates and B factors.

  • TLS parameters held in TLSIN/TLSOUT files.

  • Analyse with TLSANL program.

     libration axes, etc and ADPs

    To be implemented:

  • Allow TLS refinement prior to or simultaneously with refinement of other parameters.


E g 1 gapdh

E.g. 1 - GAPDH

  • Glyceraldehyde-3-phosphate dehydrogenase from Sulfolobus solfataricus (M.N.Isupov et al, JMB, in press)

  • P41212, 2.0Å, 2 molecules in a.s.u., each molecule has NAD-binding and catalytic domains.


E g 1 gapdh1

E.g. 1 - GAPDH

ScalingBisoTLS groupsR factorRfree

IsotropicRefined023.329.6

AnisotropicRefined 022.628.9

IsotropicRefined 121.026.4

IsotropicRefined 220.926.2

IsotropicRefined 420.826.1

Isotropic35Ų029.534.4

Isotropic35Ų126.632.1

Isotropic35Ų224.229.0

Isotropic35Ų423.828.4


E g 1 axes of libration

E.g.1: axes of libration

Refined Bs.

Blue - chain O, NAD-binding domain

Red - chain O, catalytic domain

Green - chain Q, NAD-binding domain

Yellow - chain Q, catalytic domain


E g 1 axes of libration1

E.g.1: axes of libration

Constant Bs.

Blue - chain O, NAD-binding domain

Red - chain O, catalytic domain

Green - chain Q, NAD-binding domain

Yellow - chain Q, catalytic domain


E g 2 adh

E.g.2: ADH

  • horse liver alcohol dehydrogenase (S.Ramaswamy et al).

  • apo form: C2221, 2.0Å, single chain in a.s.u.

  • DYNDOM results from apo vs. holo forms.

    ScalingBisoTLS groupsR factorRfree

    IsotropicRefined027.932.5

    AnisotropicRefined 023.629.1

    IsotropicRefined 122.527.4

    IsotropicRefined 422.327.4

    IsotropicRefined 622.227.6


E g 2 dynamic domains

E.g.2: dynamic domains

Results from DYNDOM.

Blue - first domain

Red - second domain

Green - hinge region


E g 2 axes of libration

E.g.2: axes of libration

TLS groups:

Blue - first dynamic domain

Red - second dynamic domain

Green - hinge region

Yellow - flexible loop


E g 3 lysozyme complex

E.g. 3: lysozyme complex

  • Hen egg white lysozyme complexed withcamelid single-chain antibody (K. Decanniere et al).

  • C2, 2.1Å, single copy in a.s.u.

    ScalingBisoTLS groupsR factorRfree

    IsotropicRefined020.224.3

    AnisotropicRefined 020.024.0

    IsotropicRefined 419.923.7

    IsotropicRefined 4*19.923.7


E g 3 axes of libration

E.g.3: axes of libration

Simple minimisation.

Blue - antibody

Red - CRD3 loop of antibody

Green - lysozyme

Yellow - lysozyme


E g 3 axes of libration1

E.g.3: axes of libration

Minimisation with TLS constrained to be positive semi-definite

Blue - antibody

Red - CRD3 loop of antibody

Green - lysozyme

Yellow - lysozyme


Acknowledgements

Acknowledgements

  • CCP4

  • BBSRC

  • Garib Murshudov (REFMAC)

  • Misha Isupov (GAPDH)

  • S Ramaswamy (ADH)

  • Klaas Decanniere (lysozyme complex)


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