FNR Fd All oxygenic photosynthetically derived reducing equivalents are utilized by combinations of a single m u l t i f u c t i o n a l e l e c t r o n c a r r i e r p r o t e i n , f e r r e d o x i n ( F d ) , a n d s e v e r a l F d - d e p e n d e n t o x i d o r e d u c t a s e s . P l a n t - t y p e F d i s a s m a l l , s o l u b l e , a c i d i c p r o t e i n d i s t r i b u t e d i n p l a n t s , a l g a e a n d c y a n o b a c t e r i a . E a c h F d - d e p e n d e n t e n z y m e i s a l s o d i s t r i b u t e d i n the same organism and functions in photosynthetic met abo lis m, suc h as Fd- NAD P + red uct ase ( FNR ), w h i c h i s i n v o l v e d i n t h e p r o c e s s o f c a r b o n a s s i m i l a t i o n ; n i t r i t e r e d u c t a s e a n d g l u t a m a t e sy nt ha se , w hi ch a re i nv ol ve d in n it ro ge n as si mi la ti on ; s u l f i t e r e d u c t a s e , w h i c h i s i n v o l v e d i n s u l f u r ass imi lat ion ; and fer red oxi n-t hio red oxi n red uct ase , which is involved in the redox regulation of several enzymes. About 20 years ago, the first structure of a plant- type Fd was reported [1]. Many biochemists have been studying the interaction sites of Fd based on Structure of the Electron Transfer Complex Between Ferredoxin and Ferredoxin-NADP + Reductase t h i s 3 - D s t r u c t u r e . A f t e r t h e c r y s t a l s t r u c t u r e o f FNR as a representative of Fd-dependent enzymes w a s r e p o r t e d i n 1 9 9 1 [ 2 ] , f u r t h e r e x p e r i m e n t s i n c l u d i n g c o m p u t e r m o d e l i n g a n d c o n t i n u o u s m u t a t i o n a l e x p e r i m e n t s o f t h i s p r o t e i n - p r o t e i n interaction have been carried out extensively. We determined the first crystal structure of the complex of Fd and FNR from maize leaf at 2.59 Å r e s o l u t i o n [ 3 ] ( F i g . 1 ) . T h e d i f f r a c t i o n d a t a w a s collected at beamline BL41XU . The redox partners a r e i n c l o s e c o n ta c t a t th e p r o s th e ti c g r o u p s , th e 2Fe-2S cluster of Fd and FAD of FNR, the shortest distance being 6.0 Å. Interaction mainly occurs by e l e c t r o s t a t i c f o r c e t h r o u g h s a l t b r i d g e s , a n d t h e interface near the prosthetic groups is hydrophobic ( Fig. 2 ). Interestingly, the structures of Fd and FNR i n t h e c o m p l e x a n d i n t h e f r e e s t a t e a l t e r i n a nu mb er of wa ys . Co ns is te nt wi th th is , we co nf ir me d F N R r e c o g n i t i o n s i t e s o n t h e F d p r o t e i n b y N M R s p e c t r o s c o p y o f t h e c o m p l e x i n s o l u t i o n . S u c h structural alteration is found at Glu 3 12 in the active site of FNR ( Fig. 3 ). We propose that this type of m o l e c u l a r c o m m u n i c a t i o n n o t o n l y d e t e r m i n e s optimal orientation of the two proteins for electron Fig. 1. Whole structure of the electron transfer complex between Fd (lower right) an d FN R (u pp er le ft ). Tw o pr os th et ic gr ou ps , FA D an d [2 Fe -2 S] cl us te r, ar e located at the interface of two proteins and shown in ball-and-stick models. 14 References [1] K. Fukuyama et al ., Nature 286 (1980) 522. [2] P.A. Karplus et al. , Science 251 (1991) 60. [3] Genji Kurisu, Masami Kusunoki, Etsuko Katoh, Toshimasa Yamazaki, Keizo Teshima, Yayoi Onda, Y o k o K i t a m a - A r i g a a n d T o s h i h a r u H a s e , N a t u r e Struct. Biol. 8 (2001) 117. tr an sf er , bu t al so co nt ri bu te s to th e mo du la ti on of the enzymatic properties of FNR. These structural a l t e r a t i o n s o f t w o p r o t e i n s a r e c o n s i s t e n t w i t h pr ev io us bi oc he mi ca l an d bi op hy si ca l re po rt s an d thought to be important for efficient electron transfer b e t w e e n t h e m . T h e 3 - D s t r u c t u r e o f t h e p h o t o s y n t h e t i c e l e c t r o n t r a n s f e r c o m p l e x i s imp ort ant for fur the r und ers tan din g of ass imi lat ory r e d u c t i o n a n d m o l e c u l a r r e c o g n i t i o n m e c h a n i s m c l o s e l y r e l a t e d t o t h e p h y s i o l o g i c a l c o n d i t i o n s o f higher plants. Genji Kurisu and Masami Kusunoki Osaka University E-mail: kurisu @ protein.osaka-u.ac.jp Fig. 2. Structure of the interface of Fd and F N R . T h e f i n a l 2 F o - F c m a p w a s d r a w n around the prosthetic groups. The vicinity of t h e r e d o x a c t i v e c e n t e r i s h y d r o p h o b i c a n d s e e m s t o b e s u i t a b l e f o r t h e d i r e c t e l e c t r o n transfer between two prosthetic groups. Fig. 3. Structural alteration of FNR induced by the complex formation with Fd. The structure around the FAD from single FNR is colored in yellow and that of the complex is in green. The si de ch ai n of E3 12 wa s mo ve d in to th e ac ti ve site upon the complex formation with Fd. E312 FAD FAD 15
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