X-RAY ABSORPTION SPECTROSCOPY OF Cu-Mo-S AND Fe-Mo-S SYSTEMS OF BIOLOGICAL RELEVANCE

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X-RAY ABSORPTION SPECTROSCOPY OF Cu-Mo-S AND Fe-Mo-S SYSTEMS OF BIOLOGICAL

RELEVANCE

J. Arber, A. Flood, C. Garner, S. Hasnain, B. Smith

To cite this version:

J. Arber, A. Flood, C. Garner, S. Hasnain, B. Smith. X-RAY ABSORPTION SPECTROSCOPY OF Cu-Mo-S AND Fe-Mo-S SYSTEMS OF BIOLOGICAL RELEVANCE. Journal de Physique Colloques, 1986, 47 (C8), pp.C8-1159-C8-1163. �10.1051/jphyscol:19868226�. �jpa-00226210�

X-RAY ABSORPTION SPECTROSCOPY OF Cu-Mo-S AND Fe-Mo-S SYSTEMS OF BIOLOGICAL RELEVANCE

3 . M . ARBER, A.C. FLOOD, C.D. GARNER, S.S. HASNAIN* and

B.E. SMITH*'

DepanhtIent of Chemistry, The University of Manchester, GB-Manchester M13 9PL, Great-Britain

' ~ a r e s b u r y Laboratory, Daresbury, GB-Warrington W A 4 4AD, Great-Britain

'"AFRC, Unit of Nitrogen Fixation, University of Sussex, GB-Brighton BNI 9RQ, Great-Britain

A b s t r a c t Systems c o n t a i n i n g e i t h e r i r o n o r copper coordinated t o sulphur and molybdenum are w e l l known t o be o f b i o l o g i c a l importance, t h e former i n t h e iron-molybdenum p r o t e i n and c o f a c t o r o f n i trogenase and t h e 1 a t t e r i n molybdenum-induced copper d e f i c i e n c y i n ruminants. X-ray absorption spectroscopy has been used t o study t h e metal K-edge EXAFS o f such systems and t o p r o v i d e new i n f o r m a t i o n about t h e i r s t r u c t u r e .

X-Ray Absorption Spectroscopy o f Systems Relevant t o Copper-Molybdenum antagonism i n Rumi nants

Analysis o f t h e Mo and Cu K X-ray absorption spectra o f mixtures o f bovine serum a1 bumin (BSA), copper sul phate and/or ammonium thiomolybdate, t o g e t h e r w i t h a s e r i e s o f c r y s t a l l o g r a p h i c a l l y c h a r a c t e r i s e d copper-sulphur-molybdenum compounds, has been c a r r i e d o u t i n order t o i n v e s t i g a t e t h e nature o f t h e type o f complex formed i n 'copper-molybdenum antagonism' i n ruminants, a sympton f i r s t noted i n t h e s c i e n t i f i c l i t e r a t u r e by Ferguson e t a l . ( 1 ) who observed t h a t c a t t l e g r a z i n g on mol ybdenum-ri ch s o i 1 s u f f e r e d from acute d i arrhoea and debi l j l a t i on. Many s t u d i e s have suggested t h a t t h e i n t e r a c t i o n o f thiomolybdate ([MoS41 ) w i t h copper i s t h e key t o t h e process (2,3) although t h e nature o f any complex formed between copper and thiomolybdate i s n o t known.

The BSA systems were prepared from (NH ) MoS , ^{CuS0 .5H} 0 and BSA (Sigma, F r a c t i o n V, f a t t y - a c i d f r e e ) as equimol a r (10md <olu?ions. %od$l compounds s t u d l e d were (NPr [(CH C H S)CuMoS I, (NPr [(C H SCu) MoS I, (Ph P ) 2 [ ( C u I ) MoS I.

(Ph P ) ~ I ( c & ~ ~ M O Z

f

~ a r ? s b u r y ~ y n c f l r o t g o n Source oper&ing a t an energy o f 2GeV w i t h an average c u r r e n t o f 1 5 0 m ~ (Mo K-edge o f BSA systems), 20mA (Cu K-edge o f BSk systems) on t n e Wiggler and 1.8GeV and 150mA f o r a l l model compound data. A double c r y s t a l Si(220) monochro- mator was used thus m i n i m i s i n g harmonic contamination, (4) ,m. f o u r scans being r e - corded f o r each sample and t h e data averaged. Data a n a l y s i s employed t h e s i n g l e - s c a t t e r i n g spherical-wave method f o r EXAFS c a l c u l a t i o n and p h a s e s h i f t s d e r i v e d from ab i n i t i o c a l c u l a t i o n s as described elsewhere. ( 5 - 7 ) . The a p p l i c a b i l i t y of t h e pha- s e s h i f t s was v e r i f i e d by t h e i n t e r p r e t a t i o n of t h e Cu and Mo K-edge EXAFS data f o r t h e model compounds p r e v i o u s l y l i s t e d u s i n g c r y s t a l l o g r a p h i c data from Nicholson (8).

The iYio K EXAFS d a t a are shown i n F i g u r e 1 t o g e t h e r w i t h t h e c a l c u l a t e d f i t s (parameters are given i n Table 1 ) . The i n t e r p r e t a t i o n s za_re c o n s i s t e n t w i t h an unchanged environment f o r t h e molybdenum i n t h e BSA/[MoS41 system, when compared t o thiomolybdate, where t h e Mo-S ^distance determined c r y s t a l 1 ographical l y (9) and by

Article published online by EDP Sciences and available at http://dx.doi.org/10.1051/jphyscol:19868226

C8-1160 JOURNAL DE PHYSIQUE

EXAFS ( 1 0 ) i s 2.18A. A d d i t i o n o f cu2+ produces changes i n t h e EXAFS t h a t suggest t h e f o r m a t i o n o f a 1 :1 Cu-MoS4 complex a l t h o u g h t h e i n t e r m e t a l l i c d i s t a n c e i s s l i g h t l y l o n g e r (2.728) t h a n t h a t o f t h e a p p r o p r i a t e model compound, where t h e c o r r e s p o n d i n g d i s t a n c e i s 2.636 2 0.001A ( 8 ) and more c l o s e l y agrees w i t h t h e Mo-Cu d i s t a n c e o f 2.71 A f o u n d i n t h e p o l y m e r i c NH4CuMoS4. ( 9 )

F i g u r e 1. EXAFS d a t a (Mo K edge) f o r ( a ) B S A / L M O S ~ I ~ - and ( b ) B S A / [ M O S ~ ] ~ - / C U ~ + ( f u l l 1 in e e x p e r i m e n t a l data;. broken 1 in e c a l c u l a t e d ) .

The f i r s t d e r i v a t i v e s o f t h e Mo K-edges o f t h e BSA systems and t h e t h i o m o l y b d a t e a n i o n and model compounds have a l s o been s t u d i e d . The edge i n f l e c t i o n weakens t h r o u g h t h e s e r i e s as t h e Cu:Mo r a t i o i n c r e a s e s i n d i c a t i n g a l o s s o f i n t e n s i t y i n t h e ' 1 s - 4 d ' t r a n s i t i o n . T h i s f e a t u r e i s enhanced i n t h e t e t r a h e d r a l l y c o o r d i n a t e d t h i o m o l y b d a t e a n i o n due t o m i x i n g o f t h e 3p l i g a n d and 4d metal atom$

o r b i t a l s . ( l l ) The Mo K-edge d e r i v @ i v e s f i t i n t o t h i s p a t t e r n , t h e BSA/IMoS 1

c l o s e l y r e s e m b l i n g t h a t o f LMoS41 alone, w h i l s t t h e s l i g h t weakening i n 4 t h e i n f l e c t i o n when cu2+ i s added i s an i n d i c a t i o n t h a t one o r l e s s copper i s b e i n g bound p e r molybdenum. Thus t h e evidence f r o m b o t h Mo K-edge and EXAFS d a t a suggests t h e f o r m a t i o n o f a 1 :1 Cu-MoS c p p l e x .

I n t h e case o f t h e B S A ~ system t h e f o r m o f t h e EXAFS i s r e m i n i s c e n t o f t h a t p r e v i o u s l y r e c o r d e d f o r t h e Cu K-edge o f s u p e r o x i d e dismutas~,(l2,13) where t h e EXAFS was s i m u l a t e d b y f o u r i m i d a z o l e groups. C o o r d i n a t i o n o f Cu t o i m i d a z o l e i s c o n s i s t e n t w i t h t h e proposed f i r s t b i n d i n g s i t e f o r t h i s m e t a l i n serum a1 bumins, (14,151 i .e. t h e t e r m i n a l Asp-Thr-Hi s sequen5T.

I n t e r p r e t a t i o n o f t h e Cu K EXAFS f o r t h e BSA/Cu / [ M O S ~ ] ~ - system ( T a b l e I ) i s c o n s i s t e n t w i t h t h a t p r e s e n t e d f o r t h e Mo K data, i . e . 3S a t 2.31A and 1Mo a t 2.744, w i t h i n t h e l i m i t s o f f 0 . 0 3 A accuracy. The c o o r d i n a t i o n o f t h r e e s u l p h u r s around t h e copper suggests t h a t t h e metal i s bound t o s u l p h u r f r o m t h e BSA as w e l l as t o two b r i d g i n g s u l p h u r s f r o m t h i o m o l y b d a t e . BSA c o n t a i n s one f r e e c y s t e i n e r e s i d u e and 34 h a l f - c y s t i n e groups ( w h i c h g i v e s t h e m o l e c u l e i t s ' l o o p e d ' s t r u c t u r e ) ( 1 6 ) b u t if copper i s b i n d i n g t o one o f t h e s e t h e n t h e metal s i t e must be moving f r o m i t s i n i t i a l p o s i t i o n i n t h e t e r m i n a l amino a c i d sequence t o another p a r t o f t h e m o l e c u l e when t h i o m o l y b d a t e i s p r e s e n t .

Thus d a t a f r o m b o t h metal edges suggest t h a t i n s o l u t i o n a 1 :1 complex i s formed between copper and t h i o m o l y b d a t e . The i n t e r m e t a l 1 i c d i s t a n c e i s 2.73 ? 0.03A and b o t h m e t a l s a r e c o o r d i n a t e d t o s u l p h u r atoms.

System K-edge atom N R / A ( ~ ) a 2 / ~ ' (b )

BSA/ I M O S ~ ] ~ - Mo S 4.0 2.19 0.006

Cu S 3.0 2.31 0.015

Mo 1.0 2.74 0.013

( a ) E r r o r s i n bond l e n g t h s are considered t o be 0.03A ( b ) Debye-Wal l e r parameter.

Fe K-Edge EXAFS o f N a t i v e and Cyanide Bound FeMoco

X-ray absorption spectra have been c o l l e c t e d f o r several samples o f t h e iron-molybdenum c o f a c t o r (FeMoco) o f n i trogenase, b o t h n a t i v e and cyanide bound.

There are no marked changes i n t h e EXAFS on cyanide binding, b u t t h e EXAFS o f b o t h types o f sample show s i g n i f i c a n t d i f f e r e n c e s from those o f Antonio e t a1. (17)

Samples were prepared f r o m K l e b s i e l l a pneumoniae, t h e c o f a c t o r being e x t r a c t e d i n t o N-methylformamide (NMF) producing a f i n a l i r o n c o n c e n t r a t i o n o f between 7.5 and 9.5mM. A f t e r l o a d i n g i n t o c e l l s t h e samples were s t o r e d i n l i q u i d n i t r o g e n . X-ray absorption s p e c t r a were recorded a t fheDaresbury Synchrotron Source o p e r a t i n g

a t 2GeV and an average c u r r e n t o f 150mA. A S i ( 1 11 ) monochromator was employed and a l i q u i d helium c r y o s t a t (Oxford Instruments CF1208) was used t o m a i n t a i n an average sample temperature o f 35K. Ten scans were recorded f o r each sample and t h e d a t a averaged; d a t a a n a l y s i s d e t a i l s are as given i n t h e previous s e c t i o n .

F i g u r e 2 shows t h e EXAFS and F o u r i e r t r a n s f o r m f o r FeMoco (sample 1 ) together w i t h a t h e o r e t i c a l f i t . There i s c l e a r evidence f o r b a c k s c a t t e r i n g from S a t 2.28 and f o r a second s h e l l o f Fe/Mo a t 2.7A. This i s c o n s i s t e n t w i t h p r e v i o u s l y published Fe K EXAFS r e s u l t s ( 1 7 ) and i m p l i e s t h a t FeMoco contains rhombs o f metal-sulphur atoms, i . e . two S b r i d g i n g Fe-Fe o r Fe-Mo. I n a d d i t i o n t h i s t y p e o f s t r u c t u r a l u n i t i s found i n both Fe-S and Fe-Mo-S c l u s t e r s (see, f u r example, H o l m ( l 8 ) ) .

F i g u r e 2. EXAFS and ~ o u r i e r t r a n s f o r m of FeMoca (sample 1 ) ( f u l l l i n e experimental data; broken 1 in e c a l c u l a t e d ) .

C8-1162 JOURNAL DE PHYSIQUE

However, t h e r e a r e two m a j o r d i f f e r e n c e s between t h e EXAFS d a t a shown here and t h a t p r e v i o u s l y p u b l i s h e d . ( 1 7 ) The f i r s t i s t h e o b s e r v a t i o n o f a s h e l l of metal atoms beyond t h e FeS Fe(Mo) u n i t i n d i c a t e d b y t h e peak i n t h e F o u r i e r t r a n s f o r m a t a p p r o x i m a t e l y 3.65A q ~ i ~ u r e 2). E i t h e r Fe o r Mo b a c k s c a t t e r i n g ( a t 3.7 and 3.58 r e s p e c t i v e l y ) can be f i t t e d ( s e e T a b 1 e 1 1 ) a l t h o u g h t h e d i f f e r e n c e s p e c t r a suggest t h a t Fe i s a more a p p r o p r i a t e c h o i c e . Nevertheless t h e f a c t t h a t we a r e c o n s i d e r i n g an average Fe environment (ENDOR and Mossbalrer experiments i n d i c a t e s i x d i s t i n c t Fe s i t e s i n two groups o f t h r e e e s s e n t i a l l y i d e n t i c a l s i t e s ( 1 9 ) ) i n t h e EXAFS may mean t h a t t h e r e i s a c e r t a i n amount o f b o t h t y p e s o f metal b a c k s c a t t e r i n g p r e s e n t . The presence o f a l o n g Fe-Fe(Mo) d i s t a n c e i n c r e a s e s t h e range o f p o s s i b l e s t r u c t u r a l models f o r FeMoco t o i n c l u d e t h e t y p e o f [Fe S61 u n i t s y n t h e s i s e d b y K a n a t z i d i s e t a l . (201, i .e. w i t h t w o t y p e s o f Fe-Fe d i s t a n t e o f 2.76 and 3.79A and Fe-S d i s t a n c e o f 2.288. Indeed an [Fe Mo S61 c o r e has a l s o been c h a r a c t e r i s e d and i s suggested as a p o s s i b l e s t e p towargs $he p r o d u c t i o n o f t h e more s t o i c h i o m e t r i c a l l y c o r r e c t [Fe6WoS I d e r i v a t i v e . (21 )

fie

FeMoco ( 1 ) Table I 1

Sample Parameters used t o s i m u l a t e EXAFS

(No.) Atom N R/A(a)

S 3.0 2.22

Fe 2.0 2.63

Mo 1 .O 2.74

Fe 1 .O 3.68

S 3.0 2.22

Fe 2.0 2.63

Mo 1 .O 2.74

Mo 1 .o 3.54

0 1 .O 2.06

S 3.0 2.21

Fe 2.0 2.63

Mo 1 .O 2.75

Fe 1 .O 3.68

S 3.0 2.22

F~MOCO+CN- Fe 2.0 2.65

( 2 ) Mo 1 .O 2.71

FeMoco ( 3 ) Fe 2.0 2.64

Mo 1 .O 2.73

( a ) E r r o r s i n bond d i s t a n c e s a r e c o n s i d e r e d t o be 5 0.03A f o r s h e l l s one and two and

2 0.05A f o r t h e l o n q e r s h e l l o f Fe atoms. ( b ) Debye-Waller parameter.

between t h e n a t i v e and cyanide bound average i r o n environment. The i n c l u s i o n o f l i g h t atom b a c k s c a t t e r i n g i s again n o t conclusive although epr r e s u l t s p o i n t t o t h e b i n d i n g o f two CN i o n s per molecule o f FeMoco.(23)

Thus t h e EXAFS r e s u l t s presented here i n d i c a t e t h a t FeMoco possesses a s t r u c t u r e which i n c l u d e s a Fe-Fe(Mo) separation o f ca. 3.65A and g i v e - no c l e a r evidence f o r t h e c o o r d i n a t i o n o f a l i g h t atom t o Fe. T h e a d d i t i o n o f C N produces no marked changes i n t h e EXAFS.

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